Neurogenesis by modulating angiotensin

ABSTRACT

The instant disclosure describes methods for treating diseases and conditions of the central and peripheral nervous system by stimulating or increasing neurogenesis. The invention includes compositions and methods based on modulation angiotensin activity to stimulate or activate the formation of new nerve cells.

RELATED APPLICATIONS

This application claims benefit of priority to U.S. ProvisionalApplications 60/746,859, filed May 9, 2006, which is incorporated byreference as if fully set forth. This application also claims benefit ofpriority to U.S. Provisional Application 60/807,594, filed Jul. 17,2006; and U.S. application Ser. No. 11/551,667, filed Oct. 20, 2006,both of which are incorporated by reference as if fully set forth.

FIELD OF THE INVENTION

The instant invention relates to methods for treating diseases andconditions of the central and peripheral nervous system by stimulatingor increasing neurogenesis via modulation of angiotensin activity. Theinvention includes methods based on the application of an agent whichmodulates angiotensin action to stimulate or activate the formation ofnew nerve cells. The angiotensin modulator can be used alone or incombination with another angiotensin modulator, a neurogenic agent,including a neurogenic sensitizing agent, or an anti-astrogenic agent.

BACKGROUND OF THE INVENTION

Neurogenesis is a vital process in the brains of animals and humans,whereby new nerve cells are continuously generated throughout the lifespan of the organism. The newly born cells are able to differentiateinto functional cells of the central nervous system and integrate intoexisting neural circuits in the brain. Neurogenesis is known to persistthroughout adulthood in two regions of the mammalian brain: thesubventricular zone (SVZ) of the lateral ventricles and the dentategyrus of the hippocampus. In these regions, multipotent neuralprogenitor cells (NPCs) continue to divide and give rise to newfunctional neurons and glial cells (for review Gage Mol Psychiatry. 2000May; 5(3):262-9). It has been shown that a variety of factors canstimulate adult hippocampal neurogenesis, e.g., adrenalectomy, voluntaryexercise, enriched environment, hippocampus dependent learning andanti-depressants (Yehuda. J Neurochem. 1989 July; 53(1):241-8, vanPraag. Proc Natl Acad Sci USA. 1999 Nov. 9; 96(23):13427-31, Brown. JEur J Neurosci. 2003 May; 17(10):2042-6, Gould. Science. 1999 Oct. 15;286(5439):548-52, Malberg. J Neurosci. 2000 Dec. 15; 20(24):9104-10,Santarelli. Science. 2003 Aug. 8; 301(5634):805-9). Other factors, suchas adrenal hormones, stress, age and drugs of abuse negatively influenceneurogenesis (Cameron. Neuroscience. 1994 July; 61(2):203-9, McEwen.Neuropsychopharmacology. 1999 October; 21(4):474-84, Kuhn. J Neurosci.1996 Mar. 15; 16(6):2027-33, Eisch. Am J Psychiatry. 2004 March;161(3):426).

Renin and angiotensin are components of the renin-angiotensin system(RAS) and the renin-angiotensin-aldosterone system (RAAS). The twosystems are commonly considered to function in regulating long-termblood pressure and blood volume in the body, with the RAAS acting inpart through the release of aldosterone from the adrenal cortex.

Both systems have renin and angiotensin in common, where reninproteolytically cleaves inactive angiotensinogen to form the decapeptideangiotensin I (AI). Angiotensin-converting enzyme (ACE) then cleaves AIto form the octapeptide angiotensin II (AII). Of the two angiotensins,AII has been observed to be more potent. AII acts as a vasoconstrictorto raise arterial blood pressure and decrease blood flow. AII also actson the adrenal cortex, which leads to the release of aldosterone. Inturn, aldosterone acts in the kidney to cause resorption of sodium andwater from urine. The result in an increase in the fluid volume ofblood.

The two systems are activated following blood loss or a drop in bloodpressure. Other components of the systems are the AII receptor(s) thatmediate AII activity. Angiotensin receptors are G protein-coupledreceptors which bind AII as a ligand. Subtypes of the receptors includeAT₁ and AT₂, both of which bind AII, and the AT₃ and AT₄ receptors.

Both the receptors and ACE have been the targets of manipulation totreat hypertension (high blood pressure) and other conditions. AIIreceptor antagonists, also referred to as angiotensin receptor blockersor ARBs, AT1-receptor antagonists, or sartans, are used to antagonizeAII activity by preventing AII interactions with AII receptor(s). ACEinhibitors are used to lower AII formation. Additional information isavailable, for example, in the review by Jackson, et al. in Goodman andGilman's The Pharmacological Basis of Therapeutics, 9th Edition, pp.733-754 (New York: McGraw-Hill, 1996).

Citation of the above documents is not intended as an admission that anyof the foregoing is pertinent prior art. All statements as to the dateor representation as to the contents of these documents is based on theinformation available to the applicant and does not constitute anyadmission as to the correctness of the dates or contents of thesedocuments.

BRIEF SUMMARY OF THE INVENTION

Disclosed herein are methods for the prophylaxis and treatment ofdiseases, conditions and injuries of the central and peripheral nervoussystems by stimulating or increasing neurogenesis. Aspects of theinvention include increasing neurogenesis in cases of a disease,disorder, or condition of the nervous system. Embodiments of theinvention include methods of treating a neurodegenerative disorder,neurological trauma including brain or central nervous system traumaand/or recovery therefrom, depression, anxiety, psychosis, learning andmemory disorders, and ischemia of the central and/or peripheral nervoussystems.

In one aspect, the invention includes methods of stimulating orincreasing neurogenesis. The neurogenesis may be at the level of a cellor tissue. The cell or tissue may be present in an animal subject or ahuman being, or alternatively be in an in vitro or ex vivo setting. Insome embodiments, neurogenesis is stimulated or increased in a neuralcell or tissue, such as that of the central or peripheral nervous systemof an animal or human being. In cases of an animal or human, the methodsmay be practiced in connection with one or more disease, disorder, orcondition of the nervous system as present in the animal or humansubject. Thus, embodiments of the invention include compositions ormethods of treating a disease, disorder, or condition by administeringone or more modulators of angiotensin activity as described herein. Insome embodiments, a modulator is an aldosterone receptor inhibitor, ACEinhibitor, or rennin inhibitor. In other embodiments, the modulator isan angiotensin receptor antagonist (also known as angiotensin receptorblockers or ARBs, AT1-receptor antagonists, or sartans). In someembodiments, antagonists of the AT₁, AT₂, AT₃, and/or AT₄ receptors areused.

In another aspect, the invention includes methods of using chemical orbiological entities as modulators of angiotensin activity to increaseneurogenesis. In some embodiments, a chemical entity used as a modulatoris a sulfhydryl-containing (or mercapto-containing) agent, such asalacepril or captopril (Capoten®) as non-limiting examples. In otherembodiments, the chemical entity is a dicarboxylate-containing agent,such as enalapril (Vasotec® or Renitec®) or enalaprilat; ramipril(Altace® or Tritace® or Ramace®); quinapril (Accupril®); perindopril(Coversyl®) or perindopril erbumine (Aceon®); and lisinopril (Lisodur®or Prinivil® or Zestril®) as non-limiting examples. In additionalembodiments, the chemical entity is a phosphonate-containing (orphosphate-containing) agent, such as fosinopril (Monopril®). Additionalexamples of a modulator of angiotensin activity include benazepril(Lotensin®), imidapril, moexipril (Univasc®), and trandolapril (Mavik®).In some embodiments, a modulator is administered in an alternative formsuch as an ester, that increases bioavailability upon oraladministration with subsequent conversion into metabolites with greateractivity.

Further embodiments include biological entities that are naturallyoccurring, such as casokinins and lactokinins (breakdown products ofcasein and whey), which may be administered as such to obviate the needfor their formation during digestion. Non-limiting embodiments ofangiotensin receptor antagonists include candesartan (Atacand® orRatacand®); eprosartan (Teveten®); irbesartan (Aprovel® or Karvea® orAvapro®); losartan (Cozaar® or Hyzaar®); olmesartan (Benicar®);telmisartan (Micardis® or Pritor®); and valsartan (Diovan®). Of coursethe invention includes the use of other modulators as well as acombination with one or more than one of the neurogenic agents describedherein, such as a PDE inhibitor.

In another aspect, the methods include identifying a patient sufferingfrom one or more diseases, disorders, or conditions, or a symptomthereof, and administering to the patient at least one modulator ofangiotensin activity as described herein. As non-limiting examples, themodulator is alacepril, candesartan, telmisartan, enalapril, lisinopril,and/or captopril. In some embodiments, the invention provides a methodincluding identification of a subject as in need of an increase inneurogenesis, and administering to the subject one or more modulators asdescribed herein. In other embodiments, the subject is a patient, suchas a human patient.

The invention further provides a method including administering one ormore modulators of angiotensin activity to a subject exhibiting theeffects of insufficient amounts of, or inadequate levels of,neurogenesis. In some embodiments, the subject may be one that has beensubjected to an agent that decreases or inhibits neurogenesis.Non-limiting examples of an inhibitor of neurogenesis includes opioidreceptor agonists, such as a mu receptor subtype agonist like morphine.In a related manner, the invention provides for administering one ormore modulators to a subject or person that will be subjected to anagent that decreases or inhibits neurogenesis. In some embodiments, thesubject or person may be one that is about to be administered morphineor other opioid receptor agonist, like another opiate, and so about tobe subject to a decrease or inhibition of neurogenesis. Non-limitingexamples include administering a modulator to a subject before,simultaneously with, or after, the subject is administered morphine orother opiate in connection with a surgical procedure.

Also disclosed are methods for preparing a population of neural stemcells suitable for transplantation, comprising culturing a population ofneural stem cells (NSCs) in vitro, and contacting the cultured neuralstem cells with at least one modulator of angiotensin activity. In someembodiments, the stem cells are prepared and then transferred to arecipient host animal or human. Non-limiting examples of preparationinclude 1) contact with a modulator until the cells have undergoneneurogenesis, such as that which is detectable by visual inspection,marker, or cell counting, or 2) contact with a modulator until the cellshave been sufficiently stimulated or induced toward or intoneurogenesis. The cells prepared in such a non-limiting manner may betransplanted to a subject, optionally with simultaneous, nearlysimultaneous, or subsequent administration of a neurogenic agent, or amodulator of angiotensin activity to the subject. While the neural stemcells may be in the form of an in vitro culture or cell line, in otherembodiments, the cells may be part of a tissue which is subsequentlytransplanted into a subject, based upon the cell morphology.

In yet another aspect, the invention includes methods of stimulating orincreasing neurogenesis in a subject by administering a modulator ofangiotensin activity. In some embodiments, the neurogenesis occurs incombination with the stimulation of angiogenesis which provides newcells with access to the circulatory system.

In another aspect, the invention includes the use of a modulator ofangiotensin activity in combination with a neurogenic agent, such as,but not limited to, a PDE inhibitor, such as to PDE3, 4, or 5; or GABAinhibitor. Moreover, certain combinations include candesartan, captoprilor telmisartan with ibudilast, enoximone, baclofen, donepezil,vardenafil, rolipram or theophylline. Furthermore, the combination ofagents may be administered in one formulation, or concurrently orsequentially in more than one formulation.

The details of additional embodiments of the invention are set forth inthe accompanying drawings and the description below. Other features,objects, and advantages of the invention will be apparent from thedrawings and detailed description, and from the claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a dose-response curve showing effect of the neurogenic agentalacepril on neuronal differentiation. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. EC₅₀ was observed at an alacepril concentration of 13 μM intest cells, compared to 4.7 μM for the positive control compound.

FIG. 2 is a dose-response curve showing effect of the neurogenic agentenalapril on neuronal differentiation. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. EC₅₀ was observed at an enalapril concentration of 0.5 μM intest cells, compared to 4.7 μM for the positive control compound.

FIG. 3 is a dose-response curve showing effect of the neurogenic agentlisinopril on neuronal differentiation. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. EC₅₀ was observed at a lisinopril concentration of 9.1 μM intest cells, compared to 4.7 μM for the positive control compound.

FIG. 4 is a dose-response curve showing effect of the neurogenic agentcaptopril on neuronal differentiation. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. EC₅₀ was observed at a captopril concentration of 3.8 μM intest cells, compared to 4.7 μM for the positive control compound.

FIG. 5 is a dose-response curve showing effect of the neurogenic agentlosartan on neuronal differentiation. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. EC₅₀ was observed at a losartan concentration of 0.9 μM intest cells, compared to 4.7 μM for the positive control compound.

FIG. 6 is a dose-response curve showing effect of the neurogenic agentbenazepril (ACE inhibitor) on neuronal differentiation of human neuralstem cells. The agent was tested in a concentration response curveranging from 0.01 uM to 31.6 uM. Data is presented as the percentage ofthe neuronal positive control, with basal media values subtracted.Benazepril promoted neuronal differentiation of human neural stem cellswith an EC₅₀ of 3.3 uM in test cells.

FIG. 7 is a dose-response curve showing effect of the neurogenic agenttrandolapril (ACE inhibitor) on neuronal differentiation of human neuralstem cells. The agent was tested in a concentration response curveranging from 0.01 uM to 31.6 uM. Data is presented as the percentage ofthe neuronal positive control, with basal media values subtracted.Trandolapril promoted neuronal differentiation of human neural stemcells with an EC₅₀ of approximately 35 uM in test cells.

FIG. 8 is a dose-response curve showing effect of the neurogenic agentcandesartan (angiotensin receptor blocker) on neuronal differentiationof human neural stem cells. The agent was tested in a concentrationresponse curve ranging from 0.01 uM to 31.6 uM. Data is presented as thepercentage of the neuronal positive control, with basal media valuessubtracted. Candesartan promoted neuronal differentiation of humanneural stem cells with an EC₅₀ of approximately 0.84 uM in test cells.

FIG. 9 is a dose-response curve showing effect of the neurogenic agenttelmisartan (angiotensin receptor blocker) on neuronal differentiationof human neural stem cells. The agent was tested in a concentrationresponse curve ranging from 0.001 uM to 3.16 uM. Data is presented asthe percentage of the neuronal positive control, with basal media valuessubtracted. Telmisartan promoted neuronal differentiation of humanneural stem cells with an EC₅₀ of approximately 0.03 uM in test cells.

FIG. 10 is a dose-response curve showing the effect of the neurogenicagents captopril (angiotensin converting enzyme, or ACE, inhibitor) andibudilast (PDE inhibitor) in combination on neuronal differentiationcompared to the effect of either agent alone. When run independently,ibudilast was tested in a concentration response curve (CRC) rangingfrom 0.01 μM to 10.0 μM and captopril was tested in a CRC ranging from0.01 μM to 31.6 μM. In combination, ibudilast was tested in a CRCranging from 0.003 μM to 10.0 μM and captopril was added at aconcentration 3.16-fold higher at each point (for example, the firstpoint in the combined curve reflects a combination of 0.003 μM ibudilastand 0.01 μM captopril). Data is presented as the percentage of theneuronal positive control, with basal media values subtracted. When usedalone, EC₅₀ was observed at a captopril concentration of 3.8 μM or anibudilast concentration of 6.2 μM in test cells. When used incombination, EC₅₀ was observed in a combination of captopril at aconcentration of 0.15 μM and ibudilast at a concentration of 0.05 μM,resulting in a synergistic combination index of 0.05.

FIG. 11 is a dose-response curve showing the effect of the neurogenicagents captopril (ACE inhibitor) and enoximone (PDE-3 inhibitor) incombination on neuronal differentiation compared to the effect of eitheragent alone. When run independently, each compound was tested in aconcentration response curve ranging from 0.01 μM to 31.6 μM. Incombination, the compounds were combined at equal concentrations at eachpoint (for example, the first point in the combined curve consisted of atest of 0.01 μM captopril and 0.01 μM enoximone). Data is presented asthe percentage of the neuronal positive control, with basal media valuessubtracted. When used alone, EC₅₀ was observed at a captoprilconcentration of 3.8 μM or an enoximone concentration of 6.8 μM in testcells. When used in combination, EC₅₀ was observed at a combination ofcaptopril and enoximone at concentrations of 1.1 μM each, resulting in asynergistic combination index of 0.5.

FIG. 12 is a dose-response curve showing the effect of the neurogenicagents candesartan (angiotensin II AT1 receptor antagonist) andibudilast (PDE inhibitor) in combination on neuronal differentiationcompared to the effect of either agent alone. When run independently,ibudilast was tested in a concentration response curve (CRC) rangingfrom 0.01 μM to 10.0 μM and candesartan was tested in a CRC ranging from0.01 μM to 31.6 μM. In combination, ibudilast was tested in a CRCranging from 0.003 μM to 10.0 μM and candesartan was added at aconcentration 3.16-fold higher at each point (for example, the firstpoint in the combined curve reflects a combination of 0.003 μM ibudilastand 0.01 μM captopril). Data is presented as the percentage of theneuronal positive control, with basal media values subtracted. When usedalone, EC₅₀ was observed at a candesartan concentration of 2.2 μM or anibudilast concentration of 6.2 μM in test cells. When used incombination, EC₅₀ was observed at the combination of candesartan at aconcentration of 0.35 μM and ibudilast at a concentration of 0.11 μM,resulting in a synergistic combination index of 0.07.

FIG. 13 is a dose-response curve showing the effect of the neurogenicagents captopril (ACE inhibitor) and baclofen (GABA agonist) incombination on neuronal differentiation compared to the effect of eitheragent alone. When run independently, each compound was tested in aconcentration response curve ranging from 0.01 μM to 31.6 μM. Incombination, the compounds were combined at equal concentrations at eachpoint (for example, the first point in the combined curve consisted of atest of 0.01 μM captopril and 0.01 μM baclofen). Data is presented asthe percentage of the neuronal positive control, with basal media valuessubtracted. When used alone, EC₅₀ was observed at a captoprilconcentration of 3.8 μM or a baclofen concentration of 3.2 μM in testcells. When used in combination, EC₅₀ was observed at a combination ofcaptopril and baclofen at concentrations of 1.3 μM each, resulting in asynergistic combination index of 0.88.

FIG. 14 is a dose-response curve showing the effect of the neurogenicagents captopril (ACE inhibitor) and donepezil (acetylcholinesteraseinhibitor) in combination on neuronal differentiation compared to theeffect of either agent alone. When run independently, each compound wastested in a concentration response curve ranging from 0.01 μM to 31.6μM. In combination, the compounds were combined at equal concentrationsat each point (for example, the first point in the combined curveconsisted of a test of 0.01 μM captopril and 0.01 μM donepezil). Data ispresented as the percentage of the neuronal positive control, with basalmedia values subtracted. When used alone, EC₅₀ was observed at acaptopril concentration of 3.8 μM or a donepezil concentration of 2.0 μMin test cells. When used in combination, EC₅₀ was observed at acombination of captopril and donepezil at concentrations of 0.16 μMeach, resulting in a synergistic combination index of 0.13.

FIG. 15 is a dose-response curve showing the effect of the neurogenicagents captopril (ACE inhibitor) and vardenafil (levitra, PDE5inhibitor) in combination on neuronal differentiation compared to theeffect of either agent alone. When run independently, each compound wastested in a concentration response curve ranging from 0.01 μM to 31.6μM. In combination, the compounds were combined at equal concentrationsat each point (for example, the first point in the combined curveconsisted of a test of 0.01 μM captopril and 0.01 μM vardenafil). Datais presented as the percentage of the neuronal positive control, withbasal media values subtracted. When used alone, EC₅₀ was observed at acaptopril concentration of 3.8 μM or a vardenafil concentration of 8.6μM in test cells. When used in combination, EC₅₀ was observed at acombination of captopril and vardenafil at concentrations of 1.6 μMeach, resulting in a synergistic combination index of 0.69.

FIG. 16 is a dose-response curve showing the effect of the neurogenicagents telmisartan (angiotensin II AT1 receptor antagonist) and rolipram(PDE4 inhibitor) in combination on neuronal differentiation compared tothe effect of either agent alone. When run independently, telmisartanwas tested in a concentration response curve (CRC) ranging from 0.001 μMto 3.16 μM and rolipram was tested in a CRC ranging from 0.01 μM to 31.6μM. In combination, telmisartan was tested in a CRC ranging from 0.001μM to 3.16 μM and rolipram was added at a concentration 10-fold higherat each point (for example, the first point in the combined curvereflects a combination of 0.001 μM telmisartan and 0.01 μM rolipram).Data is presented as the percentage of the neuronal positive control,with basal media values subtracted. When used alone, EC₅₀ was observedat a telmisartan concentration of 0.06 μM or a rolipram concentration of2.3 μM in test cells. When used in combination, EC₅₀ was observed at aconcentration of 0.027 μM telmisartan and 0.27 μM rolipram, resulting ina synergistic combination index of 0.62.

FIG. 17 is a dose-response curve showing the effect of the neurogenicagents captopril (ACE inhibitor) and theophylline (PDE inhibitor) incombination on neuronal differentiation compared to the effect of eitheragent alone. When run independently, each compound was tested in aconcentration response curve ranging from 0.01 μM to 31.6 μM. Incombination, the compounds were combined at equal concentrations at eachpoint (for example, the first point in the combined curve consisted of atest of 0.01 μM captopril and 0.01 μM theophylline). Data is presentedas the percentage of the neuronal positive control, with basal mediavalues subtracted. When used alone, EC₅₀ was observed at a captoprilconcentration of 3.8 μM or a theophylline concentration of 16.4 μM intest cells. When used in combination, EC₅₀ was observed at a combinationof captopril and donepezil at concentrations of 0.0.22 μM each,resulting in a synergistic combination index of 0.07.

DETAILED DESCRIPTION OF MODES OF PRACTICING THE INVENTION

“Neurogenesis” is defined herein as proliferation, differentiation,migration and/or survival of a neural cell in vivo or in vitro. Invarious embodiments, the neural cell is an adult, fetal, or embryonicneural stem cell or population of cells. The cells may be located in thecentral nervous system or elsewhere in an animal or human being. Thecells may also be in a tissue, such as neural tissue. In someembodiments, the neural cell is an adult, fetal, or embryonic progenitorcell or population of cells, or a population of cells comprising amixture of stem cells and progenitor cells. Neural cells include allbrain stem cells, all brain progenitor cells, and all brain precursorcells. Neurogenesis includes neurogenesis as it occurs during normaldevelopment, as well as neural regeneration that occurs followingdisease, damage or therapeutic intervention, such as by the treatmentdescribed herein.

A “neurogenic agent” is defined as a chemical or biological agent orreagent that can promote, stimulate, or otherwise increase the amount ordegree or nature of neurogenesis in vivo or ex vivo or in vitro relativeto the amount, degree, or nature of neurogenesis in the absence of theagent or reagent. In some embodiments, treatment with a neurogenic agentincreases neurogenesis if it promotes neurogenesis by at least about 5%,at least about 10%, at least about 25%, at least about 50%, at leastabout 100%, at least about 500%, or more in comparison to the amount,degree, and/or nature of neurogenesis in the absence of the agent, underthe conditions of the method used to detect or determine neurogenesis.As described herein, a neurogenic agent is a modulator of angiotensinactivity, such as an ACE inhibitor or angiotensin receptor antagonist.

The term “astrogenic” is defined in relation to “astrogenesis” whichrefers to the activation, proliferation, differentiation, migrationand/or survival of an astrocytic cell in vivo or in vitro. Non-limitingexamples of astrocytic cells include astrocytes, activated microglialcells, astrocyte precursors and potentiated cells, and astrocyteprogenitor and derived cells. In some embodiments, the astrocyte is anadult, fetal, or embryonic astrocyte or population of astrocytes. Theastrocytes may be located in the central nervous system or elsewhere inan animal or human being. The astrocytes may also be in a tissue, suchas neural tissue. In some embodiments, the astrocyte is an adult, fetal,or embryonic progenitor cell or population of cells, or a population ofcells comprising a mixture of stem and/or progenitor cells, that is/arecapable of developing into astrocytes. Astrogenesis includes theproliferation and/or differentiation of astrocytes as it occurs duringnormal development, as well as astrogenesis that occurs followingdisease, damage or therapeutic intervention.

The term “stem cell” (or neural stem cell (NSC)), as used herein, refersto an undifferentiated cell that is capable of self-renewal anddifferentiation into neurons, astrocytes, and/or oligodendrocytes.

The term “progenitor cell” (e.g., neural progenitor cell), as usedherein, refers to a cell derived from a stem cell that is not itself astem cell. Some progenitor cells can produce progeny that are capable ofdifferentiating into more than one cell type.

An “angiotensin receptor antagonist” is a ligand that binds theangiotensin receptor and has AII receptor antagonist activity that maybe greater than, or similar to, antagonist activity at other AIIreceptor subtypes. Non-limiting examples of receptor subtypes includeAT₁ and AT₂, both of which bind AII, as well as AT₃ and AT₄. In someembodiments, antagonist activity at one AII receptor subtype may beapproximately equal to antagonist activity at another AII receptorsubtype. In other embodiments, the antagonist activity at an AIIreceptor subtype is “selective” by being at least about 5%, at leastabout 10%, at least about 15%, at least about 20%, at least about 30%,at least about 50%, at least about 75%, at least about 100%, at leastabout 200%, at least about 300%, at least about 400%, or at least about500% or more than antagonist activity at another receptor subtype.Alternatively, antagonist activity relative to AII may be the same atone receptor subtype as at one or more other subtypes. Antagonists thatlack agonist activity at any of the AII receptor subtypes may beadvantageously used in the practice of the invention.

The present invention includes compositions and methods of increasingneurogenesis by contacting cells with one or more modulators ofangiotensin activity. The amount of a modulator of the invention, suchas an ACE inhibitor, may be selected to be effective to produce animprovement in a treated subject, or detectable neurogenesis in vitro.In some embodiments, the amount is one that also minimizes clinical sideeffects seen with administration of the agent to a subject. The amountof a modulator used in vivo may be about 50%, about 45%, about 40%,about 35%, about 30%, about 25%, about 20%, about 18%, about 16%, about14%, about 12%, about 10%, about 8%, about 6%, about 4%, about 2%, orabout 1% or less of the maximum tolerated dose for a subject. This isreadily determined for each modulator that has been in clinical use ortesting, such as in humans.

In another aspect, the invention includes compositions and methods ofusing one or more AII receptor antagonists, at a level at whichneurogenesis occur. The amount of antagonist may be any that iseffective to produce neurogenesis. In methods of increasing neurogenesisby contacting cells with an AII receptor antagonist, the cells may be invitro or in vivo. In some embodiments, the cells are present in a tissueor organ of a subject animal or human being. The AII receptor antagonistmay be any that has AII receptor selective antagonist activity asdescribed herein. The cells are those capable of neurogenesis, such asto result, whether by direct differentiation or by proliferation anddifferentiation, in differentiated neuronal or glial cells.Representative, and non-limiting examples of other AII receptorantagonist compounds for use in the present invention are providedbelow.

In applications to an animal or human being, the invention relates to amethod of bringing cells into contact with a modulator of angiotensinactivity in effective amounts to result in an increase in neurogenesisin comparison to the absence of the modulator. A non-limiting example isin the administration of the modulator to the animal or human being.Such contacting or administration may also be described as exogenouslysupplying the modulator to a cell or tissue.

In some embodiments, the term “animal” or “animal subject” refers to anon-human mammal, such as a primate, canine, or feline. In otherembodiments, the terms refer to an animal that is domesticated (e.g.livestock) or otherwise subject to human care and/or maintenance (e.g.zoo animals and other animals for exhibition). In other non-limitingexamples, the terms refer to ruminants or carnivores, such as dogs,cats, birds, horses, cattle, sheep, goats, marine animals and mammals,penguins, deer, elk, and foxes.

The present invention also relates to methods of treating diseases,disorders, and conditions of the central and/or peripheral nervoussystems (CNS and PNS, respectively) by administering one or moremodulators of angiotensin activity. As used herein, “treating” includesprevention, amelioration, alleviation, and/or elimination of thedisease, disorder, or condition being treated or one or more symptoms ofthe disease, disorder, or condition being treated, as well asimprovement in the overall well being of a patient, as measured byobjective and/or subjective criteria. In some embodiments, treating isused for reversing, attenuating, minimizing, suppressing, or haltingundesirable or deleterious effects of, or effects from the progressionof, a disease, disorder, or condition of the central and/or peripheralnervous systems. In other embodiments, the method of treating may beadvantageously used in cases where additional neurogenesis wouldreplace, replenish, or increase the numbers of cells lost due to injuryor disease as non-limiting examples.

The amount of the modulator of angiotensin activity may be any thatresults in a measurable relief of a disease condition like thosedescribed herein. As a non-limiting example, an improvement in theHamilton depression scale (HAM-D) score for depression may be used todetermine (such as quantitatively) or detect (such as qualitatively) ameasurable level of improvement in the depression of a subject.

Non-limiting examples of symptoms that may be treated with the methodsdescribed herein include abnormal behavior, abnormal movement,hyperactivity, hallucinations, acute delusions, combativeness,hostility, negativism, withdrawal, seclusion, memory defects, sensorydefects, cognitive defects, and tension. Non-limiting examples ofabnormal behavior include irritability, poor impulse control,distractibility, and aggressiveness.

In additional embodiments, an angiotensin agent as used herein includesa neurogenesis modulating agent, as defined herein, that elicits anobservable neurogenic response by producing, generating, stabilizing, orincreasing the retention of an intermediate agent which, when contactedwith an angiotensin agent, results in the neurogenic response. As usedherein, “increasing the retention of” or variants of that phrase or theterm “retention” refer to decreasing the degradation of, or increasingthe stability of, an intermediate agent.

In some cases, an angiotensin agent, optionally in combination with oneor more other neurogenic agents, results in improved efficacy, fewerside effects, lower effective dosages, less frequent dosing, and/orother desirable effects relative to use of the neurogenesis modulatingagents individually (such as at higher doses), due, e.g., to synergisticactivities and/or the targeting of molecules and/or activities that aredifferentially expressed in particular tissues and/or cell-types.

The term “neurogenic combination of an angiotensin agent with one ormore other neurogenic agents” refers to a combination of neurogenesismodulating agents. In some embodiments, administering a neurogenic, orneuromodulating, combination according to methods provided hereinmodulates neurogenesis in a target tissue and/or cell-type by at leastabout 20%, about 25%, about 30%, about 40%, about 50%, at least about75%, or at least about 90% or more in comparison to the absence of thecombination. In further embodiments, neurogenesis is modulated by atleast about 95% or by at least about 99% or more.

A neuromodulating combination may be used to inhibit a neural cell'sproliferation, division, or progress through the cell cycle.Alternatively, a neuromodulating combination may be used to stimulatesurvival and/or differentiation in a neural cell. As an additionalalternative, a neuromodulating combination may be used to inhibit,reduce, or prevent astrocyte activation and/or astrogenesis or astrocytedifferentiation.

“IC₅₀” and “EC₅₀” values are concentrations of an agent, in acombination of an angiotensin agent with one or more other neurogenicagents, that reduce and promote, respectively, neurogenesis or anotherphysiological activity (e.g., the activity of a receptor) to ahalf-maximal level. IC₅₀ and EC₅₀ values can be assayed in a variety ofenvironments, including cell-free environments, cellular environments(e.g., cell culture assays), multicellular environments (e.g., intissues or other multicellular structures), and/or in vivo. In someembodiments, one or more neurogenesis modulating agents in a combinationor method disclosed herein individually have IC₅₀ or EC₅₀ values of lessthan about 10 μM, less than about 1 μM, or less than about 0.1 μM orlower. In other embodiments, an agent in a combination has an IC₅₀ orEC₅₀ of less than about 50 nM, less than about 10 nM, less than about 1nM, less than about 0.1 nM, or lower.

In some embodiments, selectivity of one or more agents, in a combinationof a an angiotensin agent with one or more other neurogenic agents, isindividually measured as the ratio of the IC₅₀ or EC₅₀ value for adesired effect (e.g., modulation of neurogenesis) relative to theIC₅₀/EC₅₀ value for an undesired effect. In some embodiments, a“selective” agent in a combination has a selectivity of less than about1:2, less than about 1:10, less than about 1:50, or less than about1:100. In some embodiments, one or more agents in a combinationindividually exhibits selective activity in one or more organs, tissues,and/or cell types relative to another organ, tissue, and/or cell type.For example, in some embodiments, an agent in a combination selectivelymodulates neurogenesis in a neurogenic region of the brain, such as thehippocampus (e.g., the dentate gyrus), the subventricular zone, and/orthe olfactory bulb.

In other embodiments, modulation by a combination of agents is in aregion containing neural cells affected by disease or injury, regioncontaining neural cells associated with disease effects or processes, orregion containing neural cells affect other event injurious to neuralcells. Non-limiting examples of such events include stroke or radiationtherapy of the region. In additional embodiments, a neuromodulatingcombination substantially modulates two or more physiological activitiesor target molecules, while being substantially inactive against one ormore other molecules and/or activities.

The term “cognitive function” refers to mental processes of an animal orhuman subject relating to information gathering and/or processing; theunderstanding, reasoning, and/or application of information and/orideas; the abstraction or specification of ideas and/or information;acts of creativity, problem-solving, and possibly intuition; and mentalprocesses such as learning, perception, and/or awareness of ideas and/orinformation. The mental processes are distinct from those of beliefs,desires, and the like. In some embodiments, cognitive function may beassessed, and thus optionally defined, via one or more tests or assaysfor cognitive function. Non-limiting examples of a test or assay forcognitive function include CANTAB (see for example Fray et al. “CANTABbattery: proposed utility in neurotoxicology.” Neurotoxicol Teratol.1996; 18(4):499-504), Stroop Test, Trail Making, Wechsler Digit Span, orthe CogState computerized cognitive test (see also Dehaene et al.“Reward-dependent learning in neuronal networks for planning anddecision making.” Prog Brain Res. 2000; 126:217-29; Iverson et al.“Interpreting change on the WAIS-III/WMS-III in clinical samples.” ArchClin Neuropsychol. 2001; 16(2):183-91; and Weaver et al. “Mild memoryimpairment in healthy older adults is distinct from normal aging.” BrainCogn. 2006; 60(2):146-55).

Angiotensin Agents

A modulator may be a rennin inhibitor, such as aliskerin.

Also provided herein, a modulator may be a sulfhydryl-containing agent,such as alacepril, captopril (Capoten®), fentiapril, pivopril,pivalopril, or zofenopril.

Alacepril (also known as1-(D-3-acetylthio-2-methylpropanoyl)-L-prolyl-L-phenylalanine or1-[(S)-3-acetylthio-2-methylpropanoyl]-L-prolyl-L-phenylalanine) isreferenced by CAS Registry Number (CAS RN) 74258-86-9. This modulator isdescribed, for example, in Onoyama et al., Clin Pharmacol Ther, 38(4):462-8 (1985)) and is represented by the following structure:

Captopril, or 1-[(2S)-3-mercapto-2-methylpropionyl]-1-proline (orD-3-mercapto-2-methylpropanoyl-L-proline or1-(2-methyl-3-sulfanyl-propanoyl)pyrrolidine-2-carboxylic acid) isreferenced by CAS RN 62571-86-2, and is also disclosed in U.S. Pat. No.4,046,889, which is hereby incorporated by reference in its entirety asif fully set forth. Captopril is represented by the following structure:

In addition to captopril, a modulator may be a substituted acylderivative of amino acids, disclosed as ACE inhibitors, in U.S. Pat.Nos. 4,129,571 and 4,154,960, which are hereby incorporated by referencein its entirety as if fully set forth.

Fentiapril, or rentiapril, is another sulfhydryl-containing modulatordisclosed herein and in Clin. Exp. Pharmacol. Physiol. 10:131 (1983),which is incorporated by reference as if fully set forth. It isreferenced by CAS RN 80830-42-8 and has a structure represented by thefollowing:

Other rentiapril isomers, represented as follows, may also be used as amodulator of angiotensin activity as disclosed herein:

Pivopril, or(S)—N-cyclopentyl-N-[3-[(2,2-dimethyl-1-oxopropyl)thio]-2-methyl-1-oxopropyl]glycine,is another a sulfhydryl-containing modulator of angiotensin activity. Itis referenced by CAS RN 81045-50-3 and discussed by Suh et al.(“Angiotensin-converting enzyme inhibitors. New orally activeantihypertensive (mercaptoalkanoyl)- and [(acylthio)alkanoyl]glycinederivatives.” J Med Chem. 28(1):57-66, 1985). Its structure isrepresented as follows:

Pivalopril, or Rhc 3659 orN-cyclopentyl-N-(3-((2,2-dimethyl-1-oxopropyl)thio)-2-methyl-1-oxypropyl)glycine,is referenced by CAS RN 76963-39-8. It has a structure represented bythe following:

Zofenopril, referenced by CAS RN 81872-10-8, is a pro-drug that isconverted to the related sulfhydryl-containing compound zofenoprilat,referenced by CAS Registry Number 75176-37-3, which is an ACE for use asdescribed herein. Studies on the conversion in humans are described byDal Bo et al. (“Assay of zofenopril and its active metabolitezofenoprilat by liquid chromatography coupled with tandem massspectrometry.” J Chromatogr B Biomed Sci Appl. 749(2):287-94, 2000). Ithas a structure represented by the following:

The metabolite zofenoprilat (CAS RN 75176-37-3) may also be used as amodulator of angiotensin activity as described herein. Its structure isrepresented as follows:

In other embodiments, the chemical entity is a dicarboxylate-containingagent, such as enalapril (Vasotec® or Renitec®) or enalaprilat; ramipril(Altace® or Tritace® or Ramace®); quinapril (Accupril®); perindopril(Coversyl®); lisinopril (Lisodur® or Prinivil® or Zestril®); benazepril;and moexipril (Univasc®) as non-limiting examples.

Enalapril, or(S)-1-[N-[1-(ethoxycarbonyl)-3-phenylpropyl]-1-alanyl]-1-proline or1-[2-(1-ethoxycarbonyl-3-phenyl-propyl)aminopropanoyl]pyrrolidine-2-carboxylicacid or enalapril maleate, is referenced by CAS RN 75847-73-3 andPatchett et al., Nature 288, 280 (1980). It is represented by thefollowing structure:

The related metabolite compound, called enalaprilat, referenced by CASRN 76420-72-9, may also be used as a modulator of angiotensin activityas disclosed herein. It has a structure represented by the following:

Ramipril, or4-[2-(1-ethoxycarbonyl-3-phenyl-propyl)aminopropanoyl]-4-azabicyclo[3.3.0]octane-3-carboxylicacid, is referenced by CAS RN 87333-19-5. It is also disclosed in U.S.Pat. No. 4,587,258, which is hereby incorporated by reference in itsentirety as if fully set forth. Its structure is represented by thefollowing:

Ramiprilat (CAS RN 87269-97-4) is the metabolite of ramipril and mayalso be used as a modulator of angiotensin activity as described herein.Its structure is represented as follows:

Quinapril, or2-[2-(1-ethoxycarbonyl-3-phenyl-propyl)aminopropanoyl]-1,2,3,4-tetrahydroisoquinoline-3-carboxylicacid, is referenced by CAS RN 85441-61-8 and disclosed in U.S. Pat. No.4,344,949 which is hereby incorporated by reference in its entirety asif fully set forth. Its structure is represented by the following:

Quinaprilat (CAS RN 85441-60-7 or 82768-85-2) is the metabolite ofquinapril and may also be used as a modulator of angiotensin activity asdescribed herein. Its structure is represented as follows:

-   -   or

Perindopril, or perindopril erbumine, is also known as1-[2-(1-ethoxycarbonylbutylamino)propanoyl]-2,3,3a,4,5,6,7,7a-octahydroindole-2-carboxylicacid. It is referenced by CAS RN 82834-16-0 and has a structurerepresented by the following:

Perindoprilat (CAS RN 95153-31-4) is the metabolite of perindopril andmay also be used as a modulator of angiotensin activity as describedherein. Its structure is represented as follows:

Lisinopril (CAS RN 76547-98-3) or (S)-1-(N(sup2)-(1-carboxy-3-phenylpropyl)-L-lysyl)-L-proline is also known as1-[6-amino-2-(1-carboxy-3-phenyl-propyl)amino-hexanoyl]pyrrolidine-2-carboxylicacid dihydrate (CAS RN 83915-83-7). Its structure, and the structure ofthe dihydrate, are represented by the following:

Benazepril, or2-[4-(1-ethoxycarbonyl-3-phenyl-propyl)amino-5-oxo-6-azabicyclo[5.4.0]undeca-7,9,11-trien-6-yl]aceticacid, is referenced by CAS RN 86541-75-5 and disclosed in U.S. Pat. No.4,410,520, which is hereby incorporated by reference in its entirety asif fully set forth. Its structure, and the structure of the dihydrate,are represented by the following:

Benazeprilat or Cgs 14831 (referenced as CAS RN 86541-78-8 or89747-91-1) is the metabolite of benazepril and may also be used as amodulator of angiotensin activity as described herein. Its structure isrepresented as follows:

Moexipril, or2-[2-[(1-ethoxycarbonyl-3-phenyl-propyl)amino]propanoyl]-6,7-dimethoxy-3,4-dihydro-1H-isoquinoline-3-carboxylicacid, is referenced by CAS RN 103775-10-6 and its structure isrepresented by the following:

Moexiprilat (CAS RN 103775-14-0) is the metabolite of moexipril and mayalso be used as a modulator of angiotensin activity as described herein.Its structure is represented as follows:

Non-limiting embodiments of angiotensin receptor antagonists includecandesartan (Atacand® or Ratacand®); eprosartan (Teveten®); irbesartan(Aprovel® or Karvea® or Avapro®); losartan (Cozaar® or Hyzaar®);olmesartan (Benicar®); telmisartan (Micardis® or Pritor®); and valsartan(Diovan®).

Candesartan, or2-ethoxy-3-[[4-[2-(2H-tetrazol-5-yl)phenyl]phenyl]methyl]-3H-benzoimidazole-4-carboxylicacid, is referenced as CAS RN 139481-59-7. The structure of candesartanis represented by the following:

Eprosartan, or4-[[2-butyl-5-(2-carboxy-3-thiophen-2-yl-prop-1-enyl)-imidazol-1-yl]methyl]benzoicacid, is referenced by CAS RN 133040-01-4 and represented by thefollowing structure:

Irbesartan, or3-butyl-2-[[4-[2-(2H-tetrazol-5-yl)phenyl]phenyl]methyl]-2,4-diazaspiro[4.4]non-3-en-1-one,is referenced by CAS RN 138402-11-6. The structure of irbesartan isrepresented by the following:

Losartan, also known as[2-butyl-5-chloro-3-[[4-[2-(2H-tetrazol-5-yl)phenyl]phenyl]methyl]-3H-imidazol-4-yl]methanolor2-butyl-4-chloro-1-[p-(O-1H-tetrazol-5-ylphenyl)benzyl]imidazole-5-methanolmonopotassium salt, is referenced by CAS RN 114798-26-4 and disclosed inU.S. Pat. No. 5,138,069, which is hereby incorporated by reference inits entirety as if fully set forth. Losartan potassium (CAS RN124750-99-8) may also be used as a modulator and described herein. Thestructure of losartan is represented by the following:

Olmesartan, or4-(1-hydroxy-1-methylethyl)-2-propyl-1-((2′-(1H-tetrazol-5-yl)(1,1′-biphenyl)-4-yl)methyl)-1H-imidazole-5-carboxylic acid, isreferenced by CAS RN 144689-24-7 and has a structure represented by thefollowing:

Olmesartan medoxomil (CAS RN 144689-63-4), metabolically converted toolmesartan via ester hydrolysis, may also be used as described herein.The structure of olmesartan medoxomil is represented by the following:

Telmisartan, or2-[4-[[4-methyl-6-(1-methylbenzoimidazol-2-yl)-2-propyl-benzoimidazol-1-yl]methyl]phenyl]benzoicacid, is referenced by CAS RN 144701-48-4 and has a structurerepresented by the following:

Valsartan, or3-methyl-2-[pentanoyl-[[4-[2-(2H-tetrazol-5-yl)phenyl]phenyl]methyl]amino]-butanoicacid, is referenced by CAS RN 137862-53-4 and disclosed in U.S. Pat. No.5,399,578, which is hereby incorporated by reference in its entirety asif fully set forth. Valsartan has a structure represented by thefollowing:

In additional embodiments, the chemical entity is aphosphonate-containing (or phosphate-containing) agent, such asfosinopril (Monopril®), fosinoprilat, fosinopril sodium (CAS RN88889-14-9), or a structurally related ACE inhibitor. Fosinopril, or4-cyclohexyl-1-[2-[(2-methyl-1-propanoyloxy-propoxy)-(4-phenylbutyl)phosphoryl]acetyl]-pyrrolidine-2-carboxylicacid, is referenced by CAS RN 98048-97-6 and disclosed in U.S. Pat. No.4,337,201, which is incorporated by reference as if fully set forth. Thestructure of fosinopril is represented by the following:

Fosinoprilat (CAS RN 95399-71-6) is the metabolite of fosinopril and mayalso be used as a modulator of angiotensin activity as described herein.Its structure is represented as follows:

Imidapril, or(S)-3-(N—((S)-1-ethoxycarbonyl-3-phenylpropyl)-L-alanyl)-1-methyl-2-oxoimidazoline-4-carboxylicacid, is another modulator of angiotensin activity for use as describedherein. It is referenced by CAS RN 89371-37-9 and has a structurerepresented by the following:

Imidaprilat (CAS RN 89371-44-8) is the metabolite of imidapril and mayalso be used as a modulator of angiotensin activity as described herein.Its structure is represented as follows:

Trandolapril, or1-[2-[(1-ethoxycarbonyl-3-phenyl-propyl)amino]propanoyl]-2,3,3a,4,5,6,7,7a-octahydroindole-2-carboxylicacid, is another modulator of angiotensin activity for use as describedherein. It is referenced by CAS RN 87679-37-6 and represented by thefollowing:

Trandolaprilat, referenced as CAS RN 87679-71-8 or 83601-86-9, is themetabolite of trandolapril and may also be used as a modulator ofangiotensin activity as described herein. Its structure is representedas the following:

General

Methods described herein can be used to treat any disease or conditionfor which it is beneficial to promote or otherwise stimulate or increaseneurogenesis. One focus of the methods described herein is to achieve atherapeutic result by stimulating or increasing neurogenesis viamodulation of angiotensin activity. Thus, certain methods describedherein can be used to treat any disease or condition susceptible totreatment by increasing neurogenesis.

In some embodiments, a disclosed method is applied to modulatingneurogenesis in vivo, in vitro, or ex vivo. In in vivo embodiments, thecells may be present in a tissue or organ of a subject animal or humanbeing. Non-limiting examples of cells include those capable ofneurogenesis, such as to result, whether by differentiation or by acombination of differentiation and proliferation, in differentiatedneural cells. As described herein, neurogenesis includes thedifferentiation of neural cells along different potential lineages. Insome embodiments, the differentiation of neural stem or progenitor cellsis along a neuronal cell lineage to produce neurons. In otherembodiments, the differentiation is along both neuronal and glial celllineages. In additional embodiments, the disclosure further includesdifferentiation along a neuronal cell lineage to the exclusion of one ormore cell types in a glial cell lineage. Non-limiting examples of glialcell types include oligodendrocytes and radial glial cells, as well asastrocytes, which have been reported as being of an “astrogliallineage”. Therefore, embodiments of the disclosure includedifferentiation along a neuronal cell lineage to the exclusion of one ormore cell types selected from oligodendrocytes, radial glial cells, andastrocytes.

In applications to an animal or human being, the disclosure includes amethod of bringing cells into contact with an angiotensin agent,optionally in combination with one or more other neurogenic agents, ineffective amounts to result in an increase in neurogenesis in comparisonto the absence of the agent or combination. A non-limiting example is inthe administration of the agent or combination to the animal or humanbeing. Such contacting or administration may also be described asexogenously supplying the combination to a cell or tissue.

Embodiments of the disclosure include a method to treat, or lessen thelevel of, a decline or impairment of cognitive function. Also includedis a method to treat a mood disorder. In additional embodiments, adisease or condition treated with a disclosed method is associated withpain and/or addiction, but in contrast to known methods, the disclosedtreatments are substantially mediated by increasing neurogenesis. As afurther non-limiting example, a method described herein may involveincreasing neurogenesis ex vivo, such that a composition containingneural stem cells, neural progenitor cells, and/or differentiated neuralcells can subsequently be administered to an individual to treat adisease or condition.

In further embodiments, methods described herein allow treatment ofdiseases characterized by pain, addiction, and/or depression by directlyreplenishing, replacing, and/or supplementing neurons and/or glialcells. In further embodiments, methods described herein enhance thegrowth and/or survival of existing neural cells, and/or slow or reversethe loss of such cells in a neurodegenerative condition.

Where a method comprises contacting a neural cell with an angiotensinagent, the result may be an increase in neurodifferentiation. The methodmay be used to potentiate a neural cell for proliferation, and thusneurogenesis, via the one or more other agents used with the angiotensinagent in combination. Thus the disclosure includes a method ofmaintaining, stabilizing, stimulating, or increasingneurodifferentiation in a cell or tissue by use of an angiotensin agent,optionally in combination with one or more other neurogenic agents thatalso increase neurodifferentiation. The method may comprise contacting acell or tissue with an angiotensin agent, optionally in combination withone or more other neurogenic agents, to maintain, stabilize, stimulate,or increase neurodifferentiation in the cell or tissue.

The disclosure also includes a method comprising contacting the cell ortissue with an angiotensin agent in combination with one or more otherneurogenic agents where the combination stimulates or increasesproliferation or cell division in a neural cell. The increase inneuroproliferation may be due to the one or more other neurogenic agentsand/or to an angiotensin agent. In some cases, a method comprising sucha combination may be used to produce neurogenesis (in this case bothneurodifferentiation and/or proliferation) in a population of neuralcells. In some embodiments, the cell or tissue is in an animal subjector a human patient as described herein. Non-limiting examples include ahuman patient treated with chemotherapy and/or radiation, or othertherapy or condition which is detrimental to cognitive function; or ahuman patient diagnosed as having epilepsy, a condition associated withepilepsy, or seizures associated with epilepsy.

Administration of an angiotensin agent, optionally in combination withone or more other neurogenic agents, may be before, after, or concurrentwith, another agent, condition, or therapy. In some embodiments, theoverall combination may be of an angiotensin agent, optionally incombination with one or more other neurogenic agents.

Uses of an Angiotensin Agent

Embodiments include a method of modulating neurogenesis by contactingone or more neural cells with one or more angiotensin agents, optionallyin combination with one or more other neurogenic agents. The amount ofan angiotensin agent, or a combination thereof with one or more otherneurogenic agents, may be selected to be effective to produce animprovement in a treated subject, or detectable neurogenesis in vitro.In some embodiments, the amount is one that also minimizes clinical sideeffects seen with administration of the inhibitor to a subject.

Cognitive Function

In other embodiments, and if compared to a reduced level of cognitivefunction, a method of the invention may be for enhancing or improvingthe reduced cognitive function in a subject or patient. The method maycomprise administering an angiotensin agent, optionally in combinationwith one or more other neurogenic agents, to a subject or patient toenhance, or improve a decline or decrease, of cognitive function due toa therapy and/or condition that reduces cognitive function. Othermethods of the disclosure include treatment to affect or maintain thecognitive function of a subject or patient. In some embodiments, themaintenance or stabilization of cognitive function may be at a level, orthereabouts, present in a subject or patient in the absence of a therapyand/or condition that reduces cognitive function. In alternativeembodiments, the maintenance or stabilization may be at a level, orthereabouts, present in a subject or patient as a result of a therapyand/or condition that reduces cognitive function.

In further embodiments, and if compared to a reduced level of cognitivefunction due to a therapy and/or condition that reduces cognitivefunction, a method of the invention may be for enhancing or improvingthe reduced cognitive function in a subject or patient. The method maycomprise administering an angiotensin agent, or a combination thereofwith one or more other neurogenic agents, to a subject or patient toenhance or improve a decline or decrease of cognitive function due tothe therapy or condition. The administering may be in combination withthe therapy or condition.

These methods optionally include assessing or measuring cognitivefunction of the subject or patient before, during, and/or afteradministration of the treatment to detect or determine the effectthereof on cognitive function. So in one embodiment, a methods maycomprise i) treating a subject or patient that has been previouslyassessed for cognitive function and ii) reassessing cognitive functionin the subject or patient during or after the course of treatment. Theassessment may measure cognitive function for comparison to a control orstandard value (or range) in subjects or patients in the absence of anangiotensin agent, or a combination thereof with one or more otherneurogenic agents. This may be used to assess the efficacy of anangiotensin agent, alone or in a combination, in alleviating thereduction in cognitive function.

Mood Disorders

In other embodiments, a disclosed method may be used to moderate oralleviate a mood disorder in a subject or patient as described herein.Thus the disclosure includes a method of treating a mood disorder insuch a subject or patient. Non-limiting examples of the method includethose comprising administering an angiotensin agent, or a combinationthereof with one or more other neurogenic agents, to a subject orpatient that is under treatment with a therapy and/or condition thatresults in a mood disorder. The administration may be with anycombination and/or amount that is effective to produce an improvement inthe mood disorder.

Representative and non-limiting mood disorders are described herein.Non-limiting examples of mood disorders include depression, anxiety,hypomania, panic attacks, excessive elation, seasonal mood (oraffective) disorder, schizophrenia and other psychoses, lissencephalysyndrome, anxiety syndromes, anxiety disorders, phobias, stress andrelated syndromes, aggression, non-senile dementia, post-paindepression, and combinations thereof.

Identification of Subjects and Patients

The disclosure includes methods comprising identification of anindividual suffering from one or more disease, disorders, or conditions,or a symptom thereof, and administering to the subject or patient anangiotensin agent, optionally in combination with one or more otherneurogenic agents, as described herein. The identification of a subjector patient as having one or more disease, disorder or condition, or asymptom thereof, may be made by a skilled practitioner using anyappropriate means known in the field.

In some embodiments, identification of a patient in need of neurogenesismodulation comprises identifying a patient who has or will be exposed toa factor or condition known to inhibit neurogenesis, including but notlimited to, stress, aging, sleep deprivation, hormonal changes (e.g.,those associated with puberty, pregnancy, or aging (e.g., menopause),lack of exercise, lack of environmental stimuli (e.g., socialisolation), diabetes and drugs of abuse (e.g., alcohol, especiallychronic use; opiates and opioids; psychostimulants). In some cases, thepatient has been identified as non-responsive to treatment with primarymedications for the condition(s) targeted for treatment (e.g.,non-responsive to antidepressants for the treatment of depression), andan angiotensin agent, optionally in combination with one or more otherneurogenic agents, is administered in a method for enhancing theresponsiveness of the patient to a co-existing or pre-existing treatmentregimen.

In other embodiments, the method or treatment comprises administering acombination of a primary medication or therapy for the condition(s)targeted for treatment and an angiotensin agent, optionally incombination with one or more other neurogenic agents. For example, inthe treatment of depression or related neuropsychiatric disorders, acombination may be administered in conjunction with, or in addition to,electroconvulsive shock treatment, a monoamine oxidase modulator, and/ora selective reuptake modulators of serotonin and/or norepinephrine.

In additional embodiments, the patient in need of neurogenesismodulation suffers from premenstrual syndrome, post-partum depression,or pregnancy-related fatigue and/or depression, and the treatmentcomprises administering a therapeutically effective amount of anangiotensin agent, optionally in combination with one or more otherneurogenic agents. Without being bound by any particular theory, andoffered to improve understanding of the invention, it is believed thatlevels of steroid hormones, such as estrogen, are increased during themenstrual cycle during and following pregnancy, and that such hormonescan exert a modulatory effect on neurogenesis.

In some embodiments, the patient is a user of a recreational drugincluding, but not limited to, alcohol, amphetamines, PCP, cocaine, andopiates. Without being bound by any particular theory, and offered toimprove understanding of the invention, it is believed that some drugsof abuse have a modulatory effect on neurogenesis, which is associatedwith depression, anxiety and other mood disorders, as well as deficitsin cognition, learning, and memory. Moreover, mood disorders arecausative/risk factors for substance abuse, and substance abuse is acommon behavioral symptom (e.g., self medicating) of mood disorders.Thus, substance abuse and mood disorders may reinforce each other,rendering patients suffering from both conditions non-responsive totreatment. Thus, in some embodiments, an angiotensin agent, optionallyin combination with one or more other neurogenic agents, to treatpatients suffering from substance abuse and/or mood disorders. Inadditional embodiments, an angiotensin agent, optionally in combinationwith one or more other neurogenic agents, can used in combination withone or more additional agents selected from an antidepressant, anantipsychotic, a mood stabilizer, or any other agent known to treat oneor more symptoms exhibited by the patient. In some embodiments, anangiotensin agent exerts a synergistic effect with the one or moreadditional agents in the treatment of substance abuse and/or mooddisorders in patients suffering from both conditions.

In further embodiments, the patient is on a co-existing and/orpre-existing treatment regimen involving administration of one or moreprescription medications having a modulatory effect on neurogenesis. Forexample, in some embodiments, the patient suffers from chronic pain andis prescribed one or more opiate/opioid medications; and/or suffers fromADD, ADHD, or a related disorder, and is prescribed a psychostimulant,such as Ritalin®, dexedrine, adderall, or a similar medication whichinhibits neurogenesis. Without being bound by any particular theory, andoffered to improve understanding of the invention, it is believed thatsuch medications can exert a modulatory effect on neurogenesis, leadingto depression, anxiety and other mood disorders, as well as deficits incognition, learning, and memory. Thus, in some preferred embodiments, anangiotensin agent, optionally in combination with one or more otherneurogenic agents, is administered to a patient who is currently or hasrecently been prescribed a medication that exerts a modulatory effect onneurogenesis, in order to treat depression, anxiety, and/or other mooddisorders, and/or to improve cognition.

In additional embodiments, the patient suffers from chronic fatiguesyndrome; a sleep disorder; lack of exercise (e.g., elderly, infirm, orphysically handicapped patients); and/or lack of environmental stimuli(e.g., social isolation); and the treatment comprises administering atherapeutically effective amount of an angiotensin agent, optionally incombination with one or more other neurogenic agents.

In more embodiments, the patient is an individual having, or who islikely to develop, a disorder relating to neural degeneration, neuraldamage and/or neural demyelination.

In further embodiments, a subject or patient includes human beings andanimals in assays for behavior linked to neurogenesis. Exemplary humanand animal assays are known to the skilled person in the field.

In yet additional embodiments, identifying a patient in need ofneurogenesis modulation comprises selecting a population orsub-population of patients, or an individual patient, that is moreamenable to treatment and/or less susceptible to side effects than otherpatients having the same disease or condition. In some embodiments,identifying a patient amenable to treatment with an angiotensin agent,optionally in combination with one or more other neurogenic agents,comprises identifying a patient who has been exposed to a factor knownto enhance neurogenesis, including but not limited to, exercise,hormones or other endogenous factors, and drugs taken as part of apre-existing treatment regimen. In some embodiments, a sub-population ofpatients is identified as being more amenable to neurogenesis modulationwith an angiotensin agent, optionally in combination with one or moreother neurogenic agents, by taking a cell or tissue sample fromprospective patients, isolating and culturing neural cells from thesample, and determining the effect of the combination on the degree ornature of neurogenesis of the cells, thereby allowing selection ofpatients for which the therapeutic agent has a substantial effect onneurogenesis. Advantageously, the selection of a patient or populationof patients in need of or amenable to treatment with an angiotensinagent, optionally in combination with one or more other neurogenicagents, of the disclosure allows more effective treatment of the diseaseor condition targeted for treatment than known methods using the same orsimilar compounds.

In some embodiments, the patient has suffered a CNS insult, such as aCNS lesion, a seizure (e.g., electroconvulsive seizure treatment;epileptic seizures), radiation, chemotherapy and/or stroke or otherischemic injury. Without being bound by any particular theory, andoffered to improve understanding of the invention, it is believed thatsome CNS insults/injuries leads to increased proliferation of neuralstem cells, but that the resulting neural cells form aberrantconnections which can lead to impaired CNS function and/or diseases,such as temporal lobe epilepsy. In other embodiments, an angiotensinagent, optionally in combination with one or more other neurogenicagents, is administered to a patient who has suffered, or is at risk ofsuffering, a CNS insult or injury to stimulate neurogenesis.Advantageously, stimulation of the differentiation of neural stem cellswith an angiotensin agent, optionally in combination with one or moreother neurogenic agents, activates signaling pathways necessary forprogenitor cells to effectively migrate and incorporate into existingneural networks or to block inappropriate proliferation.

Methods for assessing the nature and/or degree of neurogenesis in vivoand in vitro, for detecting changes in the nature and/or degree ofneurogenesis, for identifying neurogenesis modulating agents, forisolating and culturing neural stem cells, and for preparing neural stemcells for transplantation or other purposes are disclosed, for example,in U.S. Provisional Application No. 60/697,905, and U.S. PublicationNos. 2005/0009742 and 2005/0009847, 20050032702, 2005/0031538,2005/0004046, 2004/0254152, 2004/0229291, and 2004/0185429, all of whichare herein incorporated by reference in their entirety.

Neurogenesis includes the differentiation of neural cells alongdifferent potential lineages. In some embodiments of the invention, thedifferentiation of neural stem or progenitor cells is along a neuronaland/or glial cell lineage, optionally to the exclusion ofdifferentiation along an astrocyte lineage.

A modulator of angiotensin activity as described herein includespharmaceutically acceptable salts, derivatives, prodrugs, andmetabolites of the modulator. Methods for preparing and administeringsalts, derivatives, prodrugs, and metabolites of various agents are wellknown in the art.

Compounds described herein that contain a chiral center include allpossible stereoisomers of the compound, including compositionscomprising the racemic mixture of the two enantiomers, as well ascompositions comprising each enantiomer individually, substantially freeof the other enantiomer. Thus, for example, contemplated herein is acomposition comprising the S enantiomer of a compound substantially freeof the R enantiomer, or the R enantiomer substantially free of the Senantiomer. If the named compound comprises more than one chiral center,the scope of the present disclosure also includes compositionscomprising mixtures of varying proportions between the diastereomers, aswell as compositions comprising one or more diastereomers substantiallyfree of one or more of the other diastereomers. By “substantially free”it is meant that the composition comprises less than 25%, 15%, 10%, 8%,5%, 3%, or less than 1% of the minor enantiomer or diastereomer(s).Methods for synthesizing, isolating, preparing, and administeringvarious stereoisomers are known in the art.

Methods described herein can be used to treat any disease or conditionfor which it is beneficial to promote or otherwise stimulate or increaseneurogenesis. One focus of the methods described herein is to achieve atherapeutic result by increasing neurogenesis. Thus, certain methodsdescribed herein can be used to treat any disease or conditionsusceptible to treatment by increasing neurogenesis.

In some embodiments, the disease or condition being treated isassociated with pain and/or addiction, but in contrast to known methods,the treatments of the invention are substantially mediated by increasingneurogenesis. For example, in some embodiments, methods described hereininvolve increasing neurogenesis ex vivo, such that a compositioncontaining neural stem cells, neural progenitor cells, and/ordifferentiated neural cells can subsequently be administered to anindividual to treat a disease or condition. In some embodiments, methodsdescribed herein allow treatment of diseases characterized by pain,addiction, and/or depression to be treated by directly replenishing,replacing, and/or supplementing neurons and/or glial cells. In furtherembodiments, methods described herein enhance the growth and/or survivalof existing neural cells, and/or slow or reverse the loss of such cellsin a neurodegenerative condition.

Examples of diseases and conditions treatable by the methods describedherein include, but are not limited to, neurodegenerative disorders,such as senile dementia, Alzheimer's Disease, memory disturbances/memoryloss, Parkinson's disease, Parkinson's disorders, Huntington's disease(Huntington's Chorea), Lou Gehrig's disease, multiple sclerosis, Pick'sdisease, Parkinsonism dementia syndrome, progressive subcorticalgliosis, progressive supranuclear palsy, thalmic degeneration syndrome,hereditary aphasia, amyotrophic lateral sclerosis, Shy-Drager syndrome,and Lewy body disease.

The invention also provides for the treatment of a nervous systemdisorder related to cellular degeneration, a psychiatric condition,cellular trauma and/or injury, or other neurologically relatedconditions. In practice, the invention may be applied to a subject orpatient afflicted with, or diagnosed with, one or more central orperipheral nervous system disorders in any combination. Diagnosis may beperformed by a skilled person in the applicable fields using known androutine methodologies which identify and/or distinguish these nervoussystem disorders from other conditions.

Non-limiting examples of nervous system disorders related to cellulardegeneration include neurodegenerative disorders, neural stem celldisorders, neural progenitor cell disorders, degenerative diseases ofthe retina, and ischemic disorders. In some embodiments, an ischemicdisorder comprises an insufficiency, or lack, of oxygen or angiogenesis,and non-limiting example include spinal ischemia, ischemic stroke,cerebral infarction, multi-infarct dementia. While these conditions maybe present individually in a subject or patient, the invention alsoprovides for the treatment of a subject or patient afflicted with, ordiagnosed with, more than one of these conditions in any combination.

Non-limiting embodiments of nervous system disorders related to apsychiatric condition include neuropsychiatric disorders and affectivedisorders. As used herein, an affective disorder refers to a disorder ofmood such as, but not limited to, depression, post-traumatic stressdisorder (PTSD), hypomania, panic attacks, excessive elation, bipolardepression, bipolar disorder (manic-depression), and seasonal mood (oraffective) disorder. Other non-limiting embodiments includeschizophrenia and other psychoses, lissencephaly syndrome, anxietysyndromes, anxiety disorders, phobias, stress and related syndromes,cognitive function disorders, aggression, drug and alcohol abuse,obsessive compulsive behavior syndromes, borderline personalitydisorder, non-senile dementia, post-pain depression, post-partumdepression, and cerebral palsy.

Examples of nervous system disorders related to cellular or tissuetrauma and/or injury include, but are not limited to, neurologicaltraumas and injuries, surgery related trauma and/or injury, retinalinjury and trauma, injury related to epilepsy, spinal cord injury, braininjury, brain surgery, trauma related brain injury, trauma related tospinal cord injury, brain injury related to cancer treatment, spinalcord injury related to cancer treatment, brain injury related toinfection, brain injury related to inflammation, spinal cord injuryrelated to infection, spinal cord injury related to inflammation, braininjury related to environmental toxin, and spinal cord injury related toenvironmental toxin.

Non-limiting examples of nervous system disorders related to otherneurologically related conditions include learning disorders, memorydisorders, age-associated memory impairment (AAMI) or age-related memoryloss, autism, attention deficit disorders, narcolepsy, sleep disorders,cognitive disorders, epilepsy, and temporal lobe epilepsy.

Additionally, the invention provides for the application of a modulatorof angiotensin activity to treat a subject or patient for a conditiondue to the anti-neurogenic effects of an opiate or opioid basedanalgesic. In some embodiments of the invention, the administration ofan opiate or opioid based analgesic, such as an opiate like morphine orother opioid receptor agonist, to a subject or patient results in adecrease in, or inhibition of, neurogenesis. The administration of amodulator of the invention in combination with an opiate or opioid basedanalgesic would reduce the anti-neurogenic effect. One non-limitingexample is administration of a modulator of the invention in combinationwith an opioid receptor agonist after surgery (such as for the treatingpost-operative pain).

So the invention includes a method of treating post operative pain in asubject or patient by combining administration of an opiate or opioidbased analgesic with a modulator of the invention. The analgesic mayhave been administered before, simultaneously with, or after amodulator. In some cases, the analgesic or opioid receptor agonist ismorphine or another opiate.

In some embodiments, methods of treatment disclosed herein comprise thestep of administering to a mammal a modulator of angiotensin activityfor a time and at a concentration sufficient to treat the conditiontargeted for treatment. Methods of the invention can be applied toindividuals having, or who are likely to develop, disorders relating toneural degeneration, neural damage and/or neural demyelination. In someembodiments, a method comprises selecting a population or sub-populationof patients, or selecting an individual patient, that is more amenableto treatment and/or less susceptible to side effects than other patientshaving the same disease or condition. For example, in some embodiments,a sub-population of patients is identified as being more amenable toneurogenesis with a modulator of angiotensin activity by taking a cellor tissue sample from prospective patients, isolating and culturingneural cells from the sample, and determining the effect of one or moremodulators on the degree or nature of neurogenesis, thereby allowingselection of patients for which one or more modulators have asubstantial effect on neurogenesis. Advantageously, the selectionstep(s) results in more effective treatment for the disease or conditionthat known methods using the same or similar compounds.

Transplantation

In other embodiments, methods described herein involve modulatingneurogenesis in vitro or ex vivo with an angiotensin agent, optionallyin combination with one or more other neurogenic agents, such that acomposition containing neural stem cells, neural progenitor cells,and/or differentiated neural cells can subsequently be administered toan individual to treat a disease or condition. In some embodiments, themethod of treatment comprises the steps of contacting a neural stem cellor progenitor cell with an angiotensin agent, optionally in combinationwith one or more other neurogenic agents, to modulate neurogenesis, andtransplanting the cells into a patient in need of treatment. Methods fortransplanting stem and progenitor cells are known in the art, and aredescribed, e.g., in U.S. Pat. Nos. 5,928,947; 5,817,773; and 5,800,539,and PCT Publication Nos. WO 01/176507 and WO 01/170243, all of which areincorporated herein by reference in their entirety. In some embodiments,methods described herein allow treatment of diseases or conditions bydirectly replenishing, replacing, and/or supplementing damaged ordysfunctional neurons. In further embodiments, methods described hereinenhance the growth and/or survival of existing neural cells, and/or slowor reverse the loss of such cells in a neurodegenerative or othercondition.

In alternative embodiments, the method of treatment comprisesidentifying, generating, and/or propagating neural cells in vitro or exvivo in contact with an angiotensin agent, optionally in combinationwith one or more other neurogenic agents, and transplanting the cellsinto a subject. In another embodiment, the method of treatment comprisesthe steps of contacting a neural stem cell of progenitor cell with anangiotensin agent, optionally in combination with one or more otherneurogenic agents, to stimulate neurogenesis or neurodifferentiation,and transplanting the cells into a patient in need of treatment. Alsodisclosed are methods for preparing a population of neural stem cellssuitable for transplantation, comprising culturing a population ofneural stem cells (NSCs) in vitro, and contacting the cultured neuralstem cells with an angiotensin agent, optionally in combination with oneor more other neurogenic agents, as described herein. The disclosurefurther includes methods of treating the diseases, disorders, andconditions described herein by transplanting such treated cells into asubject or patient.

In other embodiments, the method of treatment comprises identifying,generating, and/or propagating neural cells ex vivo in contact with oneor more modulators of angiotensin activity and transplanting the cellsinto a subject. In another embodiment, the method of treatment comprisesthe steps of contacting a neural stem cell of progenitor cell with oneor more modulators to stimulate neurogenesis, and transplanting thecells into a patient in need of treatment. Also disclosed are methodsfor preparing a population of neural stem cells suitable fortransplantation, comprising culturing a population of neural stem cells(NSCs) in vitro, and contacting the cultured neural stem cells with atleast one modulator described herein. The invention further includesmethods of treating the diseases, disorders, and conditions describedherein by transplanting such cells into a subject or patient.

Methods described herein may comprise administering to the subject aneffective amount of a modulator compound or pharmaceutical compositionthereof. In general, an effective amount of modulator compound(s)according to the invention is an amount sufficient, when used asdescribed herein, to stimulate or increase neurogenesis in the subjecttargeted for treatment when compared to the absence of the compound. Aneffective amount of a composition may vary based on a variety offactors, including but not limited to, the activity of the activecompound(s), the physiological characteristics of the subject, thenature of the condition to be treated, and the route and/or method ofadministration. The methods of the invention typically involve theadministration of an agent of the invention in a dosage range of 0.001ng/kg/day to 500 ng/kg/day, preferably in a dosage range of 0.05 to 200ng/kg/day. Advantageously, methods described herein allow treatment ofindications with reductions in side effects, dosage levels, dosagefrequency, treatment duration, tolerability, and/or other factors.

In some embodiments of the methods described herein, the use of amodulator of angiotensin activity having selective activity may alloweffective treatment with substantially fewer and/or less severe sideeffects compared to existing treatments. For example, AII receptorantagonists with selectivity within the CNS, can reduce side effectsassociated with activity at receptors outside the intended targettissue/organ. Established methods of treating various conditions of theCNS and PNS with compounds having activity against opioid receptors havebeen known to cause side effects including, but not limited to,sweating, diarrhea, flushing, hypotension, bradycardia,bronchoconstriction, urinary bladder contraction, nausea, vomiting,parkinsonism, and increased mortality risk. In some embodiments, methodsdescribed herein allow treatment of certain conditions with doses thatminimize these side effects.

Depending on the desired clinical result, the disclosed modulators orpharmaceutical compositions are administered by any means suitable forachieving a desired effect. Various delivery methods are known in theart and can be used to deliver a modulator to a subject or to NSCs orprogenitor cells within a tissue of interest. The delivery method willdepend on factors such as the tissue of interest, the nature of thecompound (e.g., its stability and ability to cross the blood-brainbarrier), and the duration of the experiment, among other factors. Forexample, an osmotic minipump can be implanted into a neurogenic region,such as the lateral ventricle. Alternatively, compounds can beadministered by direct injection into the cerebrospinal fluid of thebrain or spinal column, or into the eye. Compounds can also beadministered into the periphery (such as by intravenous or subcutaneousinjection, or oral delivery), and subsequently cross the blood-brainbarrier.

In various embodiments, the modulators and pharmaceutical compositionsof the invention are administered in a manner that allows them tocontact the subventricular zone (SVZ) of the lateral ventricles and/orthe dentate gyrus of the hippocampus. Examples of routes ofadministration include parenteral, e.g., intravenous, intradermal,subcutaneous, oral (e.g., inhalation), transdermal (topical),transmucosal, and rectal administration. Intranasal administrationgenerally includes, but is not limited to, inhalation of aerosolsuspensions for delivery of compositions to the nasal mucosa, tracheaand bronchioli.

In some embodiments, the disclosed combinations are administered so asto either pass through or by-pass the blood-brain barrier. Methods forallowing factors to pass through the blood-brain barrier are known inthe art, and include minimizing the size of the factor, providinghydrophobic factors which facilitate passage, and conjugating amodulator of the invention to a carrier molecule that has substantialpermeability across the blood brain barrier. In some instances, thecombination of compounds can be administered by a surgical procedureimplanting a catheter coupled to a pump device. The pump device can alsobe implanted or be extracorporally positioned. Administration of themodulator can be in intermittent pulses or as a continuous infusion.Devices for injection to discrete areas of the brain are known in theart. In certain embodiments, the modulator is administered locally tothe ventricle of the brain, substantia nigra, striatum, locus ceruleous,nucleus basalis Meynert, pedunculopontine nucleus, cerebral cortex,and/or spinal cord by, e.g., injection. Methods, compositions, anddevices for delivering therapeutics, including therapeutics for thetreatment of diseases and conditions of the CNS and PNS, are known inthe art.

In some embodiments, the delivery or targeting of an AChE inhibitor,optionally in combination with another AChE inhibitor and/or anotherneurogenic agent, to a neurogenic region, such as the dentate gyrus orthe subventricular zone, enhances efficacy and reduces side effectscompared to known methods involving administration with the same orsimilar compounds.

In embodiments to treat subjects and patients, the methods includeidentifying a patient suffering from one or more disease, disorders, orconditions, or a symptom thereof, and administering to the subject orpatient an AChE inhibitor, optionally in combination with another AChEinhibitor and/or another neurogenic agent, as described herein. Theidentification of a subject or patient as having one or more disease,disorder or condition, or a symptom thereof, may be made by a skilledpractitioner using any appropriate means known in the field.

In further embodiments, the methods may be used to treat a cell, tissue,or subject which is exhibiting decreased neurogenesis or increasedneurodegeneration. In some cases, the cell, tissue, or subject is, orhas been, subjected to, or contacted with, an agent that decreases orinhibits neurogenesis. One non-limiting example is a human subject thathas been administered morphine or other agent which decreases orinhibits neurogenesis. Non-limiting examples of other agents includeopiates and opioid receptor agonists, such as mu receptor subtypeagonists, that inhibit or decrease neurogenesis.

Thus in additional embodiments, the methods may be used to treatsubjects having, or diagnosed with, depression or other withdrawalsymptoms from morphine or other agents which decrease or inhibitneurogenesis. This is distinct from the treatment of subjects having, ordiagnosed with, depression independent of an opiate, such as that of apsychiatric nature, as disclosed herein. In other embodiments, themethods may be used to treat a subject with one or more chemicaladdiction or dependency, such as with morphine or other opiates, wherethe addiction or dependency is ameliorated or alleviated by an increasein neurogenesis.

In embodiments comprising treatment of depression, the methods mayoptionally further comprise use of one or more anti-depressant agents.Thus in the treatment of depression in a subject or patient, a methodmay comprise treatment with one or more anti-depressant agents as knownto the skilled person. Non-limiting examples of anti-depressant agentsinclude an SSRI, such as fluoxetine (Prozac®), citalopram, escitalopram,fluvoxamine, paroxetine (Paxil®), and sertraline (Zoloft®) as well asthe active ingredients of known medications including Luvox® andSerozone®; selective norepinephrine reuptake inhibitors (SNRI) such asreboxetine (Edronax®) and atomoxetine (Strattera®); selective serotonin& norepinephrine reuptake inhibitor (SSNRI) such as venlafaxine(Effexor) and duloxetine (Cymbalta); and agents like baclofen,dehydroepiandrosterone (DHEA), and DHEA sulfate (DHEAS).

The combination therapy may be of one of the above with an AChEinhibitor, optionally in combination with another AChE inhibitor and/oranother neurogenic agent, as described herein to improve the conditionof the subject or patient. Non-limiting examples of combination therapyinclude the use of lower dosages of the above which reduce side effectsof the anti-depressant agent when used alone. For example, ananti-depressant agent like fluoxetine or paroxetine or sertraline may beadministered at a reduced or limited dose, optionally also reduced infrequency of administration, in combination with an AChE inhibitor. Thereduced dose mediates a sufficient anti-depressant effect so that theside effects of the anti-depressant agent alone are reduced oreliminated.

In embodiments for treating weight gain and/or to induce weight loss, anAChE inhibitor, optionally in combination with another AChE inhibitorand/or another neurogenic agent, may be used in combination with anotheragent for treating weight gain and/or inducing weight loss. Non-limitingexamples of another agent for treating weight gain and/or inducingweight loss include various diet pills that are commercially available.

The disclosed embodiments include combination therapy, where an AChEinhibitor and one or more other compounds are used together to produceneurogenesis. When administered as a combination, the therapeuticcompounds can be formulated as separate compositions that areadministered at the same time or sequentially at different times, or thetherapeutic compounds can be given as a single composition. Theinvention is not limited in the sequence of administration.

Instead, the invention includes methods wherein treatment with an ACHEinhibitor and another neurogenic agent occurs over a period of more thanabout 48 hours, more than about 72 hours, more than about 96 hours, morethan about 120 hours, more than about 144 hours, more than about 7 days,more than about 9 days, more than about 11 days, more than about 14days, more than about 21 days, more than about 28 days, more than about35 days, more than about 42 days, more than about 49 days, more thanabout 56 days, more than about 63 days, more than about 70 days, morethan about 77 days, more than about 12 weeks, more than about 16 weeks,more than about 20 weeks, or more than about 24 weeks or more. In someembodiments, treatment by administering an ACHE inhibitor occurs atleast about 12 hours, such as at least about 24, or at least about 36hours, before administration of another neurogenic agent. Followingadministration of an AChE inhibitor, further administrations may be ofonly the other neurogenic agent in some embodiments. In otherembodiments, the first administration may be of another neurogenicagent, such as a non-AChE inhibitor neurogenic agent, and furtheradministrations may be of only an AChE inhibitor.

Opiate or Opioid Based Analgesic

Additionally, the disclosed methods provide for the application of anangiotensin agent, optionally in combination with one or more otherneurogenic agents, to treat a subject or patient for a condition due tothe anti-neurogenic effects of an opiate or opioid based analgesic. Insome embodiments, the administration of an opiate or opioid basedanalgesic, such as an opiate like morphine or other opioid receptoragonist, to a subject or patient results in a decrease in, or inhibitionof, neurogenesis. The administration of an angiotensin agent, optionallyin combination with one or more other neurogenic agents, with an opiateor opioid based analgesic would reduce the anti-neurogenic effect. Onenon-limiting example is administration of such a combination with anopioid receptor agonist after surgery (such as for the treatingpost-operative pain).

Also the disclosed embodiments include a method of treating postoperative pain in a subject or patient by combining administration of anopiate or opioid based analgesic with an angiotensin agent, optionallyin combination with one or more other neurogenic agents. The analgesicmay have been administered before, simultaneously with, or after thecombination. In some cases, the analgesic or opioid receptor agonist ismorphine or another opiate.

Other disclosed embodiments include a method to treat or preventdecreases in, or inhibition of, neurogenesis in other cases involvinguse of an opioid receptor agonist. The methods comprise theadministration of an angiotensin agent, optionally in combination withone or more other neurogenic agents, as described herein. Non-limitingexamples include cases involving an opioid receptor agonist, whichdecreases or inhibits neurogenesis, and drug addiction, drugrehabilitation, and/or prevention of relapse into addiction. In someembodiments, the opioid receptor agonist is morphine, opium or anotheropiate.

In further embodiments, the disclosure includes methods to treat a cell,tissue, or subject which is exhibiting decreased neurogenesis orincreased neurodegeneration. In some cases, the cell, tissue, or subjectis, or has been, subjected to, or contacted with, an agent thatdecreases or inhibits neurogenesis. One non-limiting example is a humansubject that has been administered morphine or other agent whichdecreases or inhibits neurogenesis. Non-limiting examples of otheragents include opiates and opioid receptor agonists, such as mu receptorsubtype agonists, that inhibit or decrease neurogenesis.

Thus in additional embodiments, the methods may be used to treatsubjects having, or diagnosed with, depression or other withdrawalsymptoms from morphine or other agents which decrease or inhibitneurogenesis. This is distinct from the treatment of subjects having, ordiagnosed with, depression independent of an opiate, such as that of apsychiatric nature, as disclosed herein. In further embodiments, themethods may be used to treat a subject with one or more chemicaladdiction or dependency, such as with morphine or other opiates, wherethe addiction or dependency is ameliorated or alleviated by an increasein neurogenesis.

In other embodiments, the neurogenic agent may be an opioid ornon-opioid (acts independently of an opioid receptor) agent. In somecases, the neurogenic agent is one that antagonizes one or more opioidreceptors or is an inverse agonist of at least one opioid receptor. Anopioid receptor antagonist or inverse agonist of the invention may bespecific or selective (or alternatively non-specific or non-selective)for opioid receptor subtypes. So an antagonist may be non-specific ornon-selective such that it antagonizes more than one of the three knownopioid receptor subtypes, identified as OP₁, OP₂, and OP₃ (also know asdelta, or δ, kappa, or κ, and mu, or μ, respectively). Thus an opioidthat antagonizes any two, or all three, of these subtypes, or an inverseagonist that is specific or selective for any two or all three of thesesubtypes, may be used as the non-AChE inhibitor neurogenic agent.Alternatively, an antagonist or inverse agonist may be specific orselective for one of the three subtypes, such as the kappa subtype as anon-limiting example.

In some embodiments, the neurogenic agent used in the methods describedherein has “selective” activity (such as in the case of an antagonist orinverse agonist) under certain conditions against one or more opioidreceptor subtypes with respect to the degree and/or nature of activityagainst one or more other opioid receptor subtypes. For example, in someembodiments, the neurogenic agent has an antagonist effect against oneor more subtypes, and a much weaker effect or substantially no effectagainst other subtypes. As another example, an additional neurogenicagent used in the methods described herein may act as an agonist at oneor more opioid receptor subtypes and as antagonist at one or more otheropioid receptor subtypes. In some embodiments, a neurogenic agent hasactivity against kappa opioid receptors, while having substantiallylesser activity against one or both of the delta and mu receptorsubtypes. In other embodiments, a neurogenic agent has activity againsttwo opioid receptor subtypes, such as the kappa and delta subtypes. Asnon-limiting examples, the agents naloxone and naltrexone havenonselective antagonist activities against more than one opioid receptorsubtypes. In certain embodiments, selective activity of one or moreopioid antagonists results in enhanced efficacy, fewer side effects,lower effective dosages, less frequent dosing, or other desirableattributes.

An opioid receptor antagonist is an agent able to inhibit one or morecharacteristic responses of an opioid receptor or receptor subtype. As anon-limiting example, an antagonist may competitively ornon-competitively bind to an opioid receptor, an agonist or partialagonist (or other ligand) of a receptor, and/or a downstream signalingmolecule to inhibit a receptor's function.

An inverse agonist able to block or inhibit a constitutive activity ofan opioid receptor may also be used. An inverse agonist maycompetitively or non-competitively bind to an opioid receptor and/or adownstream signaling molecule to inhibit a receptor's function.Non-limiting examples of inverse agonists for use in the practice of theinvention include ICI-174864 (N,N-diallyl-Tyr-Aib-Aib-Phe-Leu),RTI-5989-1, RTI-5989-23, and RTI-5989-25 (see Zaki et al. J. Pharmacol.Exp. Therap. 298(3): 1015-1020, 2001).

Neurogenesis with Angiogenesis

In additional embodiments, the disclosure includes a method ofstimulating or increasing neurogenesis in a subject or patient withstimulation of angiogenesis in the subject or patient. Theco-stimulation may be used to provide the differentiating and/orproliferating cells with increased access to the circulatory system. Theneurogenesis is produced by modulation of angiotensin activity, such aswith an angiotensin agent, optionally in combination with one or moreother neurogenic agents, as described herein. An increase inangiogenesis may be mediated by a means known to the skilled person,including administration of a angiogenic factor or treatment with anangiogenic therapy. Non-limiting examples of angiogenic factors orconditions include vascular endothelial growth factor (VEGF),angiopoietin-1 or -2, erythropoietin, exercise, or a combinationthereof.

So in some embodiments, the disclosure includes a method comprisingadministering i) an angiotensin agent, optionally in combination withone or more other neurogenic agents, and ii) one or more angiogenicfactors to a subject or patient. In other embodiments, the disclosureincludes a method comprising administering i) an angiotensin agent,optionally in combination with one or more other neurogenic agents, to asubject or patient with ii) treating said subject or patient with one ormore angiogenic conditions. The subject or patient may be any asdescribed herein.

The co-treatment of a subject or patient includes simultaneous treatmentor sequential treatment as non-limiting examples. In cases of sequentialtreatment, the administration of an angiotensin agent, optionally withone or more other neurogenic agents, may be before or after theadministration of an angiogenic factor or condition. Of course in thecase of a combination of an angiotensin agent and one or more otherneurogenic agents, the angiotensin agent may be administered separatelyfrom the one or more other agents, such that the one or more other agentis administered before or after administration of an angiogenic factoror condition.

Additional Diseases and Conditions

As described herein, the disclosed embodiments include methods oftreating diseases, disorders, and conditions of the central and/orperipheral nervous systems (CNS and PNS, respectively) by administeringan angiotensin agent, optionally in combination with one or more otherneurogenic agents. As used herein, “treating” includes prevention,amelioration, alleviation, and/or elimination of the disease, disorder,or condition being treated or one or more symptoms of the disease,disorder, or condition being treated, as well as improvement in theoverall well being of a patient, as measured by objective and/orsubjective criteria. In some embodiments, treating is used forreversing, attenuating, minimizing, suppressing, or halting undesirableor deleterious effects of, or effects from the progression of, adisease, disorder, or condition of the central and/or peripheral nervoussystems. In other embodiments, the method of treating may beadvantageously used in cases where additional neurogenesis wouldreplace, replenish, or increase the numbers of cells lost due to injuryor disease as non-limiting examples.

The amount of an angiotensin agent, optionally in combination with oneor more other neurogenic agents may be any that results in a measurablerelief of a disease condition like those described herein. As anon-limiting example, an improvement in the Hamilton depression scale(HAM-D) score for depression may be used to determine (such asquantitatively) or detect (such as qualitatively) a measurable level ofimprovement in the depression of a subject.

Non-limiting examples of symptoms that may be treated with the methodsdescribed herein include abnormal behavior, abnormal movement,hyperactivity, hallucinations, acute delusions, combativeness,hostility, negativism, withdrawal, seclusion, memory defects, sensorydefects, cognitive defects, and tension. Non-limiting examples ofabnormal behavior include irritability, poor impulse control,distractibility, and aggressiveness. Outcomes from treatment with thedisclosed methods include improvements in cognitive function orcapability in comparison to the absence of treatment.

Additional examples of diseases and conditions treatable by the methodsdescribed herein include, but are not limited to, neurodegenerativedisorders and neural disease, such as dementias (e.g., senile dementia,memory disturbances/memory loss, dementias caused by neurodegenerativedisorders (e.g., Alzheimer's, Parkinson's disease, Parkinson'sdisorders, Huntington's disease (Huntington's Chorea), Lou Gehrig'sdisease, multiple sclerosis, Pick's disease, Parkinsonism dementiasyndrome), progressive subcortical gliosis, progressive supranuclearpalsy, thalmic degeneration syndrome, hereditary aphasia, amyotrophiclateral sclerosis, Shy-Drager syndrome, and Lewy body disease; vascularconditions (e.g., infarcts, hemorrhage, cardiac disorders); mixedvascular and Alzheimer's; bacterial meningitis; Creutzfeld-JacobDisease; and Cushing's disease).

The disclosed embodiments also provide for the treatment of a nervoussystem disorder related to neural damage, cellular degeneration, apsychiatric condition, cellular (neurological) trauma and/or injury(e.g., subdural hematoma or traumatic brain injury), toxic chemicals(e.g., heavy metals, alcohol, some medications), CNS hypoxia, or otherneurologically related conditions. In practice, the disclosedcompositions and methods may be applied to a subject or patientafflicted with, or diagnosed with, one or more central or peripheralnervous system disorders in any combination. Diagnosis may be performedby a skilled person in the applicable fields using known and routinemethodologies which identify and/or distinguish these nervous systemdisorders from other conditions.

Non-limiting examples of nervous system disorders related to cellulardegeneration include neurodegenerative disorders, neural stem celldisorders, neural progenitor cell disorders, degenerative diseases ofthe retina, and ischemic disorders. In some embodiments, an ischemicdisorder comprises an insufficiency, or lack, of oxygen or angiogenesis,and non-limiting example include spinal ischemia, ischemic stroke,cerebral infarction, multi-infarct dementia. While these conditions maybe present individually in a subject or patient, the disclosed methodsalso provide for the treatment of a subject or patient afflicted with,or diagnosed with, more than one of these conditions in any combination.

Non-limiting embodiments of nervous system disorders related to apsychiatric condition include neuropsychiatric disorders and affectivedisorders. As used herein, an affective disorder refers to a disorder ofmood such as, but not limited to, depression, post-traumatic stressdisorder (PTSD), hypomania, panic attacks, excessive elation, bipolardepression, bipolar disorder (manic-depression), and seasonal mood (oraffective) disorder. Other non-limiting embodiments includeschizophrenia and other psychoses, lissencephaly syndrome, anxietysyndromes, anxiety disorders, phobias, stress and related syndromes(e.g., panic disorder, phobias, adjustment disorders, migraines),cognitive function disorders, aggression, drug and alcohol abuse, drugaddiction, and drug-induced neurological damage, obsessive compulsivebehavior syndromes, borderline personality disorder, non-seniledementia, post-pain depression, post-partum depression, and cerebralpalsy.

Examples of nervous system disorders related to cellular or tissuetrauma and/or injury include, but are not limited to, neurologicaltraumas and injuries, surgery related trauma and/or injury, retinalinjury and trauma, injury related to epilepsy, cord injury, spinal cordinjury, brain injury, brain surgery, trauma related brain injury, traumarelated to spinal cord injury, brain injury related to cancer treatment,spinal cord injury related to cancer treatment, brain injury related toinfection, brain injury related to inflammation, spinal cord injuryrelated to infection, spinal cord injury related to inflammation, braininjury related to environmental toxin, and spinal cord injury related toenvironmental toxin.

Non-limiting examples of nervous system disorders related to otherneurologically related conditions include learning disorders, memorydisorders, age-associated memory impairment (AAMI) or age-related memoryloss, autism, learning or attention deficit disorders (ADD or attentiondeficit hyperactivity disorder, ADHD), narcolepsy, sleep disorders andsleep deprivation (e.g., insomnia, chronic fatigue syndrome), cognitivedisorders, epilepsy, injury related to epilepsy, and temporal lobeepilepsy.

Other non-limiting examples of diseases and conditions treatable by themethods described herein include, but are not limited to, hormonalchanges (e.g., depression and other mood disorders associated withpuberty, pregnancy, or aging (e.g., menopause)); and lack of exercise(e.g., depression or other mental disorders in elderly, paralyzed, orphysically handicapped patients); infections (e.g., HIV); geneticabnormalities (down syndrome); metabolic abnormalities (e.g., vitaminB12 or folate deficiency); hydrocephalus; memory loss separate fromdementia, including mild cognitive impairment (MCI), age-relatedcognitive decline, and memory loss resulting from the use of generalanesthetics, chemotherapy, radiation treatment, post-surgical trauma, ortherapeutic intervention; and diseases of the of the peripheral nervoussystem (PNS), including but not limited to, PNS neuropathies (e.g.,vascular neuropathies, diabetic neuropathies, amyloid neuropathies, andthe like), neuralgias, neoplasms, myelin-related diseases, etc.

Other conditions that can be beneficially treated by increasingneurogenesis are known in the art (see e.g., U.S. Publication Nos.20020106731, 2005/0009742 and 2005/0009847, 20050032702, 2005/0031538,2005/0004046, 2004/0254152, 2004/0229291, and 2004/0185429, hereinincorporated by reference in their entirety).

Formulations and Doses

In some embodiments of the disclosure, an angiotensin agent, optionallyin combination with another angiotensin agent or one or more otherneurogenic agents, is in the form of a single or multiple compositionsthat includes at least one pharmaceutically acceptable excipient. Asused herein, the term “pharmaceutically acceptable excipient” includesany excipient known in the field as suitable for pharmaceuticalapplication. Suitable pharmaceutical excipients and formulations areknown in the art and are described, for example, in Remington'sPharmaceutical Sciences (19th ed.) (Genarro, ed. (1995) Mack PublishingCo., Easton, Pa.). Preferably, pharmaceutical carriers are chosen basedupon the intended mode of administration of an angiotensin agent,optionally in combination with one or more other neurogenic agents. Thepharmaceutically acceptable carrier may include, for example,disintegrants, binders, lubricants, glidants, emollients, humectants,thickeners, silicones, flavoring agents, and water.

An angiotensin agent, optionally in combination with one or more otherneurogenic agents, or with another angiotensin agent, may beincorporated with excipients and administered in the form of ingestibletablets, buccal tablets, troches, capsules, elixirs, suspensions,syrups, wafers, or any other form known in the pharmaceutical arts. Thepharmaceutical compositions may also be formulated in a sustainedrelease form. Sustained release compositions, enteric coatings, and thelike are known in the art. Alternatively, the compositions may be aquick release formulation.

The amount of a combination of an angiotensin agent, or a combinationthereof with one or more other neurogenic agents, may be an amount thatalso potentiates or sensitizes, such as by activating or inducing cellsto differentiate, a population of neural cells for neurogenesis. Thedegree of potentiation or sensitization for neurogenesis may bedetermined with use of the combination in any appropriate neurogenesisassay, including, but not limited to, a neuronal differentiation assaydescribed herein. In some embodiments, the amount of a combination of anangiotensin agent, optionally in combination with one or more otherneurogenic agents, is based on the highest amount of one agent in acombination, which amount produces no detectable neuroproliferation invitro but yet produces neurogenesis, or a measurable shift in efficacyin promoting neurogenesis in vitro, when used in the combination.

As disclosed herein, an effective amount of an angiotensin agent,optionally in combination with one or more other neurogenic agents, inthe described methods is an amount sufficient, when used as describedherein, to stimulate or increase neurogenesis in the subject targetedfor treatment when compared to the absence of the combination. Aneffective amount of an angiotensin agent alone or in combination mayvary based on a variety of factors, including but not limited to, theactivity of the active compounds, the physiological characteristics ofthe subject, the nature of the condition to be treated, and the routeand/or method of administration. General dosage ranges of certaincompounds are provided herein and in the cited references based onanimal models of CNS diseases and conditions. Various conversionfactors, formulas, and methods for determining human dose equivalents ofanimal dosages are known in the art, and are described, e.g., inFreireich et al., Cancer Chemother Repts 50(4): 219 (1966), Monro etal., Toxicology Pathology, 23: 187-98 (1995), Boxenbaum and Dilea, J.Clin. Pharmacol. 35: 957-966 (1995), and Voisin et al., Reg. Toxicol.Pharmacol., 12(2): 107-116 (1990), which are herein incorporated byreference.

The disclosed methods typically involve the administration of anangiotensin agent, optionally in combination with one or more otherneurogenic agents, in a dosage range of from about 0.001 ng/kg/day toabout 200 mg/kg/day. Other non-limiting dosages include from about 0.001to about 0.01 ng/kg/day, about 0.01 to about 0.1 ng/kg/day, about 0.1 toabout 1 ng/kg/day, about 1 to about 10 ng/kg/day, about 10 to about 100ng/kg/day, about 100 ng/kg/day to about 1 μg/kg/day, about 1 to about 2μg/kg/day, about 2 μg/kg/day to about 0.02 mg/kg/day, about 0.02 toabout 0.2 mg/kg/day, about 0.2 to about 2 mg/kg/day, about 2 to about 20mg/kg/day, or about 20 to about 200 mg/kg/day. However, as understood bythose skilled in the art, the exact dosage of an angiotensin agent,optionally in combination with one or more other neurogenic agents, usedto treat a particular condition will vary in practice due to a widevariety of factors. Accordingly, dosage guidelines provided herein arenot limiting as the range of actual dosages, but rather provide guidanceto skilled practitioners in selecting dosages useful in the empiricaldetermination of dosages for individual patients. Advantageously,methods described herein allow treatment of one or more conditions withreductions in side effects, dosage levels, dosage frequency, treatmentduration, safety, tolerability, and/or other factors. So where suitabledosages for an angiotensin agent are known to a skilled person, thedisclosure includes the use of about 75%, about 50%, about 33%, about25%, about 20%, about 15%, about 10%, about 5%, about 2.5%, about 1%,about 0.5%, about 0.25%, about 0.2%, about 0.1%, about 0.05%, about0.025%, about 0.02%, about 0.01%, or less than the known dosage.

In other embodiments, the amount of an angiotensin agent used in vivomay be about 50%, about 45%, about 40%, about 35%, about 30%, about 25%,about 20%, about 18%, about 16%, about 14%, about 12%, about 10%, about8%, about 6%, about 4%, about 2%, or about 1% or less than the maximumtolerated dose for a subject, including where one or more otherneurogenic agents is used in combination with an angiotensin agent. Thisis readily determined for each muscarinic agent that has been inclinical use or testing, such as in humans.

Alternatively, the amount of an angiotensin agent, optionally incombination with one or more other neurogenic agents, may be an amountselected to be effective to produce an improvement in a treated subjectbased on detectable neurogenesis in vitro as described above. In someembodiments, such as in the case of a known angiotensin agent, theamount is one that minimizes clinical side effects seen withadministration of the agent to a subject. The amount of an agent used invivo may be about 50%, about 45%, about 40%, about 35%, about 30%, about25%, about 20%, about 18%, about 16%, about 14%, about 12%, about 10%,about 8%, about 6%, about 4%, about 2%, or about 1% or less of themaximum tolerated dose in terms of acceptable side effects for asubject. This is readily determined for each angiotensin agent or otheragent(s) of a combination disclosed herein as well as those that havebeen in clinical use or testing, such as in humans.

In other embodiments, the amount of an additional neurogenic sensitizingagent in a combination with an angiotensin agent of the disclosure isthe highest amount which produces no detectable neurogenesis when thesensitizing agent is used, alone in vitro, or in vivo, but yet producesneurogenesis, or a measurable shift in efficacy in promotingneurogenesis, when used in combination with an angiotensin agent.Embodiments include amounts which produce about 1%, about 2%, about 4%,about 6%, about 8%, about 10%, about 12%, about 14%, about 16%, about18%, about 20%, about 25%, about 30%, about 35%, or about 40% or more ofthe neurogenesis seen with the amount that produces the highest level ofneurogenesis in an in vitro assay.

In some embodiments, the amount may be the lowest needed to produce adesired, or minimum, level of detectable neurogenesis or beneficialeffect. Of course the administered angiotensin agent, alone or in acombination disclosed herein, may be in the form of a pharmaceuticalcomposition.

As described herein, the amount of an angiotensin agent, optionally incombination with one or more other neurogenic agents, may be any that iseffective to produce neurogenesis, optionally with reduced or minimizedamounts of astrogenesis. As a non-limiting example described herein, thelevels of astrogenesis observed with the use of certain angiotensinagents alone may be reduced or suppressed when an angiotensin agent isused in combination with a second agent such as baclofen (or other GABAmodulator with the same anti-astrogenesis activity) or melatonin. Thisbeneficial effect is observed along with the ability of each combinationof agents to stimulate neurogenesis. So while certain angiotensin agentshave been observed to produce astrogenesis, their use with a secondcompound, such as baclofen and melatonin, advantageously provides ameans to suppress the overall level of astrogenesis.

Therefore, the methods of the disclosure further include a method ofdecreasing the level of astrogenesis in a cell or cell population bycontacting the cell or population with an angiotensin agent and a secondagent that reduces or suppresses the amount or level of astrogenesiscaused by said angiotensin agent. The reduction or suppression ofastrogenesis may be readily determined relative to the amount or levelof astrogenesis in the absence of the second agent. In some embodiments,the second agent is baclofen or melatonin.

In some embodiments, an effective, neurogenesis modulating amount of acombination of an angiotensin agent, optionally in combination with oneor more other neurogenic agents, is an amount of an angiotensin agent(or of each agent in a combination) that achieves a concentration withinthe target tissue, using the particular mode of administration, at orabove the IC₅₀ or EC₅₀ for activity of target molecule or physiologicalprocess. In some cases, an angiotensin agent, optionally in combinationwith one or more other neurogenic agents, is administered in a mannerand dosage that gives a peak concentration of about 1, about 1.5, about2, about 2.5, about 5, about 10, about 20 or more times the IC₅₀ or EC₅₀concentration of an angiotensin agent (or each agent in thecombination). IC₅₀ and EC₅₀ values and bioavailability data for anangiotensin agent and other agent(s) described herein are known in theart, and are described, e.g., in the references cited herein or can bereadily determined using established methods. In addition, methods fordetermining the concentration of a free compound in plasma andextracellular fluids in the CNS, as well pharmacokinetic properties, areknown in the art, and are described, e.g., in de Lange et al., AAPSJournal, 7(3): 532-543 (2005). In some embodiments, an angiotensinagent, optionally in combination with one or more other neurogenicagents, described herein is administered, as a combination or separateagents used together, at a frequency of at least about once daily, orabout twice daily, or about three or more times daily, and for aduration of at least about 3 days, about 5 days, about 7 days, about 10days, about 14 days, or about 21 days, or about 4 weeks, or about 2months, or about 4 months, or about 6 months, or about 8 months, orabout 10 months, or about 1 year, or about 2 years, or about 4 years, orabout 6 years or longer.

In other embodiments, an effective, neurogenesis modulating amount is adose that produces a concentration of an angiotensin agent (or eachagent in a combination) in an organ, tissue, cell, and/or other regionof interest that includes the ED₅₀ (the pharmacologically effective dosein 50% of subjects) with little or no toxicity. IC₅₀ and EC₅₀ values forthe modulation of neurogenesis can be determined using methods describedin PCT Application US06/026677, filed Jul. 7, 2006, incorporated byreference, or by other methods known in the art. In some embodiments,the IC₅₀ or EC₅₀ concentration for the modulation of neurogenesis issubstantially lower than the IC₅₀ or EC₅₀ concentration for activity ofan angiotensin agent and/or other agent(s) at non-targeted moleculesand/or physiological processes.

In some methods described herein, the application of an angiotensinagent in combination with one or more other neurogenic agents may alloweffective treatment with substantially fewer and/or less severe sideeffects compared to existing treatments. In some embodiments,combination therapy with an angiotensin agent and one or more additionalneurogenic agents allows the combination to be administered at dosagesthat would be sub-therapeutic when administered individually or whencompared to other treatments. In other embodiments, each agent in acombination of agents may be present in an amount that results in fewerand/or less severe side effects than that which occurs with a largeramount. Thus the combined effect of the neurogenic agents will provide adesired neurogenic activity while exhibiting fewer and/or less severeside effects overall. In further embodiments, methods described hereinallow treatment of certain conditions for which treatment with the sameor similar compounds is ineffective using known methods due, forexample, to dose-limiting side effects, toxicity, and/or other factors.

Routes of Administration

As described, the methods of the disclosure comprise contacting a cellwith an angiotensin agent, optionally in combination with one or moreother neurogenic agents, or administering such an agent or combinationto a subject, to result in neurogenesis. Some embodiments comprise theuse of one angiotensin agent, such as buspirone, tandospirone,azasetron, granisetron, ondansetron, mosapride, cisapride, orsumatriptan, in combination with one or more other neurogenic agents. Inother embodiments, a combination of two or more agents, such as two ormore of buspirone, tandospirone, azasetron, granisetron, ondansetron,mosapride, cisapride, and sumatriptan, is used in combination with oneor more other neurogenic agents.

In some embodiments, methods of treatment disclosed herein comprise thestep of administering to a mammal an angiotensin agent, optionally incombination with one or more other neurogenic agents, for a time and ata concentration sufficient to treat the condition targeted fortreatment. The disclosed methods can be applied to individuals having,or who are likely to develop, disorders relating to neural degeneration,neural damage and/or neural demyelination.

Depending on the desired clinical result, the disclosed agents orpharmaceutical compositions are administered by any means suitable forachieving a desired effect. Various delivery methods are known in theart and can be used to deliver an agent to a subject or to NSCs orprogenitor cells within a tissue of interest. The delivery method willdepend on factors such as the tissue of interest, the nature of thecompound (e.g., its stability and ability to cross the blood-brainbarrier), and the duration of the experiment or treatment, among otherfactors. For example, an osmotic minipump can be implanted into aneurogenic region, such as the lateral ventricle. Alternatively,compounds can be administered by direct injection into the cerebrospinalfluid of the brain or spinal column, or into the eye. Compounds can alsobe administered into the periphery (such as by intravenous orsubcutaneous injection, or oral delivery), and subsequently cross theblood-brain barrier.

In some embodiments, the disclosed agents or pharmaceutical compositionsare administered in a manner that allows them to contact thesubventricular zone (SVZ) of the lateral ventricles and/or the dentategyrus of the hippocampus. The delivery or targeting of an angiotensinagent, optionally in combination with one or more other neurogenicagents, to a neurogenic region, such as the dentate gyrus or thesubventricular zone, may enhances efficacy and reduces side effectscompared to known methods involving administration with the same orsimilar compounds. Examples of routes of administration includeparenteral, e.g., intravenous, intradermal, subcutaneous, oral (e.g.,inhalation), transdermal (topical), transmucosal, and rectaladministration. Intranasal administration generally includes, but is notlimited to, inhalation of aerosol suspensions for delivery ofcompositions to the nasal mucosa, trachea and bronchioli.

In other embodiments, a combination of an angiotensin agent, optionallyin combination with one or more other neurogenic agents, is administeredso as to either pass through or by-pass the blood-brain barrier. Methodsfor allowing factors to pass through the blood-brain barrier are knownin the art, and include minimizing the size of the factor, providinghydrophobic factors which facilitate passage, and conjugation to acarrier molecule that has substantial permeability across the bloodbrain barrier. In some instances, an agent or combination of agents canbe administered by a surgical procedure implanting a catheter coupled toa pump device. The pump device can also be implanted or beextracorporally positioned. Administration of an angiotensin agent,optionally in combination with one or more other neurogenic agents, canbe in intermittent pulses or as a continuous infusion. Devices forinjection to discrete areas of the brain are known in the art. Incertain embodiments, the combination is administered locally to theventricle of the brain, substantia nigra, striatum, locus ceruleous,nucleus basalis of Meynert, pedunculopontine nucleus, cerebral cortex,and/or spinal cord by, e.g., injection. Methods, compositions, anddevices for delivering therapeutics, including therapeutics for thetreatment of diseases and conditions of the CNS and PNS, are known inthe art.

In some embodiments, an angiotensin agent and/or other agent(s) in acombination is modified to facilitate crossing of the gut epithelium.For example, in some embodiments, an angiotensin agent or other agent(s)is a prodrug that is actively transported across the intestinalepithelium and metabolized into the active agent in systemic circulationand/or in the CNS.

In other embodiments, an angiotensin agent and/or other agent(s) of acombination is conjugated to a targeting domain to form a chimerictherapeutic, where the targeting domain facilitates passage of theblood-brain barrier (as described above) and/or binds one or moremolecular targets in the CNS. In some embodiments, the targeting domainbinds a target that is differentially expressed or displayed on, or inclose proximity to, tissues, organs, and/or cells of interest. In somecases, the target is preferentially distributed in a neurogenic regionof the brain, such as the dentate gyrus and/or the SVZ. For example, insome embodiments, an angiotensin agent and/or other agent(s) of acombination is conjugated or complexed with the fatty aciddocosahexaenoic acid (DHA), which is readily transported across theblood brain barrier and imported into cells of the CNS.

Representative Conditions and Agents

The disclosure includes methods for treating depression and otherneurological diseases and conditions. In some embodiments, a method maycomprise use of a combination of an angiotensin agent and one or moreagents reported as anti-depressant agents. Thus a method may comprisetreatment with an angiotensin agent and one or more reportedanti-depressant agents as known to the skilled person. Non-limitingexamples of such agents include an SSRI (selective serotonine reuptakeinhibitor), such as fluoxetine (Prozac®; described, e.g., in U.S. Pat.Nos. 4,314,081 and 4,194,009), citalopram (Celexa®; described, e.g., inU.S. Pat. No. 4,136,193), escitalopram (Lexapro®; described, e.g., inU.S. Pat. No. 4,136,193), fluvoxamine (described, e.g., in U.S. Pat. No.4,085,225) or fluvoxamine maleate (CAS RN: 61718-82-9) and Luvox®,paroxetine (Paxil®; described, e.g., in U.S. Pat. Nos. 3,912,743 and4,007,196), or sertraline (Zoloft®; described, e.g., in U.S. Pat. No.4,536,518), or alaproclate; the compound nefazodone (Serozone®;described, e.g., in U.S. Pat. No. 4,338,317); a selective norepinephrinereuptake inhibitor (SNRI) such as reboxetine (Edronax®), atomoxetine(Strattera®), milnacipran (described, e.g., in U.S. Pat. No. 4,478,836),sibutramine or its primary amine metabolite (BTS 54 505), amoxapine, ormaprotiline; a selective serotonin and norepinephrine reuptake inhibitor(SSNRI) such as venlafaxine (Effexor®; described, e.g., in U.S. Pat. No.4,761,501), and its reported metabolite desvenlafaxine, or duloxetine(Cymbalta®; described, e.g., in U.S. Pat. No. 4,956,388); a serotonin,noradrenaline, and dopamine “triple uptake inhibitor”, such as

DOV 102,677 (see Popik et al. “Pharmacological Profile of the “Triple”Monoamine Neurotransmitter Uptake Inhibitor, DOV 102,677.” Cell Mol.Neurobiol. 2006 Apr. 25; Epub ahead of print),

DOV 216,303 (see Beer et al. “DOV 216,303, a “triple” reuptakeinhibitor: safety, tolerability, and pharmacokinetic profile.” J ClinPharmacol. 2004 44(12): 1360-7),

DOV 21,947 ((+)-1-(3,4-dichlorophenyl)-3-azabicyclo-(3.1.0)hexanehydrochloride), see Skolnick et al. “Antidepressant-like actions of DOV21,947: a “triple” reuptake inhibitor.” Eur J Pharmacol. 2003461(2-3):99-104),

NS-2330 or tesofensine (CAS RN 402856-42-2), or NS 2359 (CAS RN843660-54-8);

and agents like dehydroepiandrosterone (DHEA), and DHEA sulfate (DHEAS),CP-122,721 (CAS RN 145742-28-5).

Additional non-limiting examples of such agents include a tricycliccompound such as clomipramine, dosulepin or dothiepin, lofepramine(described, e.g., in U.S. Pat. No. 4,172,074), trimipramine,protriptyline, amitriptyline, desipramine (described, e.g., in U.S. Pat.No. 3,454,554), doxepin, imipramine, or nortriptyline; a psychostimulantsuch as dextroamphetamine and methylphenidate; an MAO inhibitor such asselegiline (Emsam®); an ampakine such as CX516 (or Ampalex®, CAS RN:154235-83-3), CX546 (or 1-(1,4-benzodioxan-6-ylcarbonyl)piperidine), andCX614 (CAS RN 191744-13-5) from Cortex Pharmaceuticals; a V1b antagonistsuch as SSR149415((2S,4R)-1-[5-chloro-1-[(2,4-dimethoxyphenyl)sulfonyl]-3-(2-methoxy-phenyl)-2-oxo-2,3-dihydro-1H-indol-3-yl]-4-hydroxy-N,N-dimethyl-2-pyrrolidinecarboxamide),

[1-(beta-mercapto-beta,beta-cyclopentamethylenepropionic acid),2-O-ethyltyrosine, 4-valine]arginine vasopressin (d(CH₂)₅[Tyr(Et₂)]VAVP(WK 1-1),

9-desglycine[1-(beta-mercapto-beta,beta-cyclopentamethylenepropionicacid), 2-O-ethyltyrosine, 4-valine]arginine vasopressindesGly9d(CH₂)₅[Tyr(Et₂)]-VAVP (WK 3-6), or

9-desglycine[1-(beta-mercapto-beta,beta-cyclopentamethylenepropionicacid),2-D-(O-ethyl)tyrosine, 4-valine]arginine vasopressin desGly9d(CH₂)₅[D-Tyr(Et₂)]VAVP (AO 3-21); a corticotropin-releasing factorreceptor (CRF) R antagonist such as CP-154,526 (structure disclosed inSchulz et al. “CP-154,526: a potent and selective nonpeptide antagonistof corticotropin releasing factor receptors.” Proc Natl Acad Sci USA.1996 93(19):10477-82), NBI 30775 (also known as R121919 or2,5-dimethyl-3-(6-dimethyl-4-methylpyridin-3-yl)-7-dipropylaminopyrazolo[1,5-a]pyrimidine),astressin (CAS RN 170809-51-5), or a photoactivatable analog thereof asdescribed in Bonk et al. “Novel high-affinity photoactivatableantagonists of corticotropin-releasing factor (CRF)” Eur. J. Biochem.267:3017-3024 (2000), or AAG561 (from Novartis); a melanin concentratinghormone (MCH) antagonist such as3,5-dimethoxy-N-(1-(naphthalen-2-ylmethyl)piperidin-4-yl)benzamide or(R)-3,5-dimethoxy-N-(1-(naphthalen-2-ylmethyl)-pyrrolidin-3-yl)benzamide(see Kim et al. “Identification of substituted 4-aminopiperidines and3-aminopyrrolidines as potent MCH-R1 antagonists for the treatment ofobesity.” Bioorg Med Chem. Lett. 2006 Jul. 29; [Epub ahead of print] forboth), or any MCH antagonist disclosed in U.S. Pat. No. 7,045,636 orpublished U.S. Patent Application US2005/0171098.

Further non-limiting examples of such agents include a tetracycliccompound such as mirtazapine (described, e.g., in U.S. Pat. No.4,062,848; see CAS RN 61337-67-5; also known as Remeron®, or CAS RN85650-52-8), mianserin (described, e.g., in U.S. Pat. No. 3,534,041), orsetiptiline.

Further non-limiting examples of such agents include agomelatine (CAS RN138112-76-2), pindolol (CAS RN 13523-86-9), antalarmin (CAS RN157284-96-3), mifepristone (CAS RN 84371-65-3), nemifitide (CAS RN173240-15-8) or nemifitide ditriflutate (CAS RN 204992-09-6), YKP-10A orR228060 (CAS RN 561069-23-6), trazodone (CAS RN 19794-93-5), bupropion(CAS RN 34841-39-9 or 34911-55-2) or bupropion hydrochloride (orWellbutrin®, CAS RN 31677-93-7) and its reported metabolite radafaxine(CAS RN 192374-14-4), NS2359 (CAS RN 843660-54-8), Org 34517 (CAS RN189035-07-2), Org 34850 (CAS RN 162607-84-3), vilazodone (CAS RN163521-12-8), CP-122,721 (CAS RN 145742-28-5), gepirone (CAS RN83928-76-1), SR58611 (see Mizuno et al. “The stimulation ofbeta(3)-adrenoceptor causes phosphorylation of extracellularsignal-regulated kinases 1 and 2 through a G(s)- but not G(i)-dependentpathway in 3T3-L1 adipocytes.” Eur J Pharmacol. 2000 404(1-2):63-8),saredutant or SR 48968 (CAS RN 142001-63-6), PRX-00023(N-{3-[4-(4-cyclohexylmethanesulfonylaminobutyl)piperazin-1-yl]phenyl}acetamide,see Becker et al. “An integrated in silico 3D model-driven discovery ofa novel, potent, and selective amidosulfonamide 5-HT1A agonist(PRX-00023) for the treatment of anxiety and depression.” J Med. Chem.2006 49(11):3116-35), vestipitant (or GW597599, CAS RN 334476-46-9),OPC-14523 or VPI-013 (see Bermack et al. “Effects of the potentialantidepressant OPC-14523[1-[3-[4-(3-chlorophenyl)-1-piperazinyl]propyl]-5-methoxy-3,4-dihydro-2-quinolinonemonomethanesulfonate] a combined sigma and 5-HT1A ligand: modulation ofneuronal activity in the dorsal raphe nucleus.” J Pharmacol Exp Ther.2004 310(2):578-83), casopitant or GW679769 (CAS RN 852393-14-7),elzasonan or CP-448,187 (CAS RN 361343-19-3), GW823296 (see publishedU.S. Patent Application US2005/0119248), delucemine or NPS 1506 (CAS RN186495-49-8), or ocinaplon (CAS RN 96604-21-6).

Yet additional non-limiting examples of such agents include CX717 fromCortex Pharmaceuticals, TGBA01AD (a serotonin reuptake inhibitor, 5-HT2agonist, 5-HT1A agonist, and 5-HT1D agonist) from Fabre-KramerPharmaceuticals, Inc., ORG 4420 (an NaSSA (noradrenergic/specificserotonergic antidepressant) from Organon, CP-316,311 (a CRF1antagonist) from Pfizer, BMS-562086 (a CRF1 antagonist) fromBristol-Myers Squibb, GW876008 (a CRF1 antagonist) fromNeurocrine/GlaxoSmithKline, ONO-2333Ms (a CRF1 antagonist) from OnoPharmaceutical Co., Ltd., JNJ-19567470 or TS-041 (a CRF1 antagonist)from Janssen (Johnson & Johnson) and Taisho, SSR 125543 or SSR 126374 (aCRF1 antagonist) from Sanofi-Aventis, Lu AA21004 and Lu AA24530 (bothfrom H. Lundbeck A/S), SEP-225289 from Sepracor Inc., ND7001 (a PDE2inhibitor) from Neuro3d, SSR 411298 or SSR 101010 (a fatty acid amidehydrolase, or FAAH, inhibitor) from Sanofi-Aventis, 163090 (a mixedserotonin receptor inhibitor) from GlaxoSmithKline, SSR 241586 (an NK2and NK3 receptor antagonist) from Sanofi-Aventis, SAR 102279 (an NK2receptor antagonist) from Sanofi-Aventis, YKP581 from SK Pharmaceuticals(Johnson & Johnson), R1576 (a GPCR modulator) from Roche, or ND1251 (aPDE4 inhibitor) from Neuro3d.

In other embodiments, a method may comprise use of a combination of anangiotensin agent and one or more agents reported as anti-psychoticagents. Non-limiting examples of a reported anti-psychotic agent as amember of a combination include olanzapine, quetiapine (Seroquel®),clozapine (CAS RN 5786-21-0) or its metabolite ACP-104(N-desmethylclozapine or norclozapine, CAS RN 6104-71-8), reserpine,aripiprazole, risperidone, ziprasidone, sertindole, trazodone,paliperidone (CAS RN 144598-75-4), mifepristone (CAS RN 84371-65-3),bifeprunox or DU-127090 (CAS RN 350992-10-8), asenapine or ORG 5222 (CASRN 65576-45-6), iloperidone (CAS RN 133454-47-4), ocaperidone (CAS RN129029-23-8), SLV 308 (CAS RN 269718-83-4), licarbazepine or GP 47779(CAS RN 29331-92-8), Org 34517 (CAS RN 189035-07-2), ORG 34850 (CAS RN162607-84-3), Org 24448 (CAS RN 211735-76-1), lurasidone (CAS RN367514-87-2), blonanserin or lonasen (CAS RN 132810-10-7), talnetant orSB-223412 (CAS RN 174636-32-9), secretin (CAS RN 1393-25-5) or humansecretin (CAS RN 108153-74-8) which are endogenous pancreatic hormones,ABT 089 (CAS RN 161417-03-4), SSR 504734 (see compound 13 in Hashimoto“Glycine Transporter Inhibitors as Therapeutic Agents forSchizophrenia.” Recent Patents on CNS Drug Discovery, 2006 1:43-53), MEM3454 (see Mazurov et al. “Selective alpha7 nicotinic acetylcholinereceptor ligands.” Curr Med Chem. 2006 13(13):1567-84), aphosphodiesterase 10A (PDE10A) inhibitor such as papaverine (CAS RN58-74-2) or papaverine hydrochloride (CAS RN 61-25-6), paliperidone (CASRN 144598-75-4), trifluoperazine (CAS RN 117-89-5), or trifluoperazinehydrochloride (CAS RN 440-17-5).

Additional non-limiting examples of such agents include trifluoperazine,fluphenazine, chlorpromazine, perphenazine, thioridazine, haloperidol,loxapine, mesoridazine, molindone, pimoxide, or thiothixene, SSR 146977(see Emonds-Alt et al. “Biochemical and pharmacological activities ofSSR 146977, a new potent nonpeptide tachykinin NK3 receptor antagonist.”Can J Physiol Pharmacol. 2002 80(5):482-8), SSR181507((3-exo)-8-benzoyl-N-[[(2s)7-chloro-2,3-dihydro-1,4-benzodioxin-1-yl]methyl]-8-azabicyclo[3.2.1]octane-3-methanaminemonohydrochloride), or SLV313(1-(2,3-dihydro-benzo[1,4]dioxin-5-yl)-4-[5-(4-fluorophenyl)-pyridin-3-ylmethyl]-piperazine).

Further non-limiting examples of such agents include Lu-35-138 (aD4/5-HT antagonist) from Lundbeck, AVE 1625 (a CB1 antagonist) fromSanofi-Aventis, SLV 310,313 (a 5-HT2A antagonist) from Solvay, SSR181507 (a D2/5-HT2 antagonist) from Sanofi-Aventis, GW07034 (a 5-HT6antagonist) or GW773812 (a D2,5-HT antagonist) from GlaxoSmithKline, YKP1538 from SK Pharmaceuticals, SSR 125047 (a sigma receptor antagonist)from Sanofi-Aventis, MEM1003 (a L-type calcium channel modulator) fromMemory Pharmaceuticals, JNJ-17305600 (a GLYT1 inhibitor) from Johnson &Johnson, XY 2401 (a glycine site specific NMDA modulator) from Xytis,PNU 170413 from Pfizer, RGH-188 (a D2, D3 antagonist) from Forrest, SSR180711 (an alpha7 nicotinic acetylcholine receptor partial agonist) orSSR 103800 (a GLYT1 (Type 1 glycine transporter) inhibitor) or SSR241586 (a NK3 antagonist) from Sanofi-Aventis.

In other disclosed embodiments, a reported anti-psychotic agent may beone used in treating schizophrenia. Non-limiting examples of a reportedanti-schizophrenia agent as a member of a combination with anangiotensin agent include molindone hydrochloride (MOBAN®) and TC-1827(see Bohme et al. “In vitro and in vivo characterization of TC-1827, anovel brain α4β2 nicotinic receptor agonist with pro-cognitiveactivity.” Drug Development Research 2004 62(1):26-40).

In some embodiments, a method may comprise use of a combination of anangiotensin agent and one or more agents reported for treating weightgain, metabolic syndrome, or obesity, and/or to induce weight loss orprevent weight gain. Non-limiting examples of the reported agent includevarious diet pills that are commercially or clinically available. Insome embodiments, the reported agent is orlistat (CAS RN 96829-58-2),sibutramine (CAS RN 106650-56-0) or sibutramine hydrochloride (CAS RN84485-00-7), phetermine (CAS RN 122-09-8) or phetermine hydrochloride(CAS RN 1197-21-3), diethylpropion or amfepramone (CAS RN 90-84-6) ordiethylpropion hydrochloride, benzphetamine (CAS RN 156-08-1) orbenzphetamine hydrochloride, phendimetrazine (CAS RN 634-03-7 or21784-30-5) or phendimetrazine hydrochloride (CAS RN 17140-98-6) orphendimetrazine tartrate, rimonabant (CAS RN 168273-06-1), bupropionhydrochloride (CAS RN: 31677-93-7), topiramate (CAS RN 97240-79-4),zonisamide (CAS RN 68291-97-4), or APD-356 (CAS RN 846589-98-8).

In other non-limiting embodiments, the agent may be fenfluramine orPondimin® (CAS RN 458-24-2), dexfenfluramine or Redux® (CAS RN3239-44-9), or levofenfluramine (CAS RN 37577-24-5); or a combinationthereof or a combination with phentermine. Non-limiting examples includea combination of fenfluramine and phentermine (or “fen-phen”) and ofdexfenfluramine and phentermine (or “dexfen-phen”).

The combination therapy may be of one of the above with an angiotensinagent as described herein to improve the condition of the subject orpatient. Non-limiting examples of combination therapy include the use oflower dosages of the above additional agents, or combinations thereof,which reduce side effects of the agent or combination when used alone.For example, an anti-depressant agent like fluoxetine or paroxetine orsertraline may be administered at a reduced or limited dose, optionallyalso reduced in frequency of administration, in combination with anangiotensin agent.

Similarly, a combination of fenfluramine and phentermine, or phentermineand dexfenfluramine, may be administered at a reduced or limited dose,optionally also reduced in frequency of administration, in combinationwith an angiotensin agent. The reduced dose or frequency may be thatwhich reduces or eliminates the side effects of the combination.

In light of the positive recitation (above and below) of combinationswith alternative agents to treat conditions disclosed herein, thedisclosure includes embodiments with the explicit exclusion of one ormore of the alternative agents or one or more types of alternativeagents. As would be recognized by the skilled person, a description ofthe whole of a plurality of alternative agents (or classes of agents)necessarily includes and describes subsets of the possible alternatives,such as the part remaining with the exclusion of one or more of thealternatives or exclusion of one or more classes.

Representative Combinations

As indicated herein, the disclosure includes combination therapy, wherean angiotensin agent in combination with one or more other neurogenicagents is used to produce neurogenesis. When administered as acombination, the therapeutic compounds can be formulated as separatecompositions that are administered at the same time or sequentially atdifferent times, or the therapeutic compounds can be given as a singlecomposition. The methods of the disclosure are not limited in thesequence of administration.

Instead, the disclosure includes methods wherein treatment with anangiotensin agent and another neurogenic agent occurs over a period ofmore than about 48 hours, more than about 72 hours, more than about 96hours, more than about 120 hours, more than about 144 hours, more thanabout 7 days, more than about 9 days, more than about 11 days, more thanabout 14 days, more than about 21 days, more than about 28 days, morethan about 35 days, more than about 42 days, more than about 49 days,more than about 56 days, more than about 63 days, more than about 70days, more than about 77 days, more than about 12 weeks, more than about16 weeks, more than about 20 weeks, or more than about 24 weeks or more.In some embodiments, treatment by administering an angiotensin agent,occurs at least about 12 hours, such as at least about 24, or at leastabout 36 hours, before administration of another neurogenic agent.Following administration of an angiotensin agent, furtheradministrations may be of only the other neurogenic agent in someembodiments of the disclosure. In other embodiments, furtheradministrations may be of only an angiotensin agent.

In some cases, combination therapy with an angiotensin agent and one ormore additional agents results in a enhanced efficacy, safety,therapeutic index, and/or tolerability, and/or reduced side effects(frequency, severity, or other aspects), dosage levels, dosagefrequency, and/or treatment duration. Examples of compounds useful incombinations described herein are provided above and below. Structures,synthetic processes, safety profiles, biological activity data, methodsfor determining biological activity, pharmaceutical preparations, andmethods of administration relating to the compounds are known in the artand/or provided in the cited references, all of which are hereinincorporated by reference in their entirety. Dosages of compoundsadministered in combination with an angiotensin agent can be, e.g., adosage within the range of pharmacological dosages established inhumans, or a dosage that is a fraction of the established human dosage,e.g., 70%, 50%, 30%, 10%, or less than the established human dosage.

In some embodiments, the neurogenic agent combined with an angiotensinagent may be a reported opioid or non-opioid (acts independently of anopioid receptor) agent. In some embodiments, the neurogenic agent is onereported as antagonizing one or more opioid receptors or as an inverseagonist of at least one opioid receptor. A opioid receptor antagonist orinverse agonist may be specific or selective (or alternativelynon-specific or non-selective) for opioid receptor subtypes. So anantagonist may be non-specific or non-selective such that it antagonizesmore than one of the three known opioid receptor subtypes, identified asOP₁, OP₂, and OP₃ (also know as delta, or δ, kappa, or κ, and mu, or μ,respectively). Thus an opioid that antagonizes any two, or all three, ofthese subtypes, or an inverse agonist that is specific or selective forany two or all three of these subtypes, may be used as the neurogenicagent in the practice. Alternatively, an antagonist or inverse agonistmay be specific or selective for one of the three subtypes, such as thekappa subtype as a non-limiting example.

Non-limiting examples of reported opioid antagonists include naltrindol,naloxone, naloxene, naltrexone, JDTic (Registry Number 785835-79-2; alsoknown as 3-isoquinolinecarboxamide,1,2,3,4-tetrahydro-7-hydroxy-N-[(1S)-1-[[(3R,4R)-4-(3-hydroxyphenyl)-3,4-dimethyl-1-piperidinyl]methyl]-2-methylpropyl]-dihydrochloride,(3R)-(9CI)), nor-binaltorphimine, and buprenorphine. In someembodiments, a reported selective kappa opioid receptor antagonistcompound, as described in US 20020132828, U.S. Pat. No. 6,559,159,and/or WO 2002/053533, may be used. All three of these documents areherein incorporated by reference in their entireties as if fully setforth. Further non-limiting examples of such reported antagonists is acompound disclosed in U.S. Pat. No. 6,900,228 (herein incorporated byreference in its entirety), arodyn (Ac[Phe(1,2,3),Arg(4),d-Ala(8)]DynA-(1-11)NH(2), as described in Bennett, et al. (2002) J. Med. Chem.45:5617-5619), and an active analog of arodyn as described in Bennett etal. (2005) J Pept Res. 65(3):322-32, alvimopan.

In some embodiments, the neurogenic agent used in the methods describedherein has “selective” activity (such as in the case of an antagonist orinverse agonist) under certain conditions against one or more opioidreceptor subtypes with respect to the degree and/or nature of activityagainst one or more other opioid receptor subtypes. For example, in someembodiments, the neurogenic agent has an antagonist effect against oneor more subtypes, and a much weaker effect or substantially no effectagainst other subtypes. As another example, an additional neurogenicagent used in the methods described herein may act as an agonist at oneor more opioid receptor subtypes and as antagonist at one or more otheropioid receptor subtypes. In some embodiments, a neurogenic agent hasactivity against kappa opioid receptors, while having substantiallylesser activity against one or both of the delta and mu receptorsubtypes. In other embodiments, a neurogenic agent has activity againsttwo opioid receptor subtypes, such as the kappa and delta subtypes. Asnon-limiting examples, the agents naloxone and naltrexone havenonselective antagonist activities against more than one opioid receptorsubtypes. In certain embodiments, selective activity of one or moreopioid antagonists results in enhanced efficacy, fewer side effects,lower effective dosages, less frequent dosing, or other desirableattributes.

An opioid receptor antagonist is an agent able to inhibit one or morecharacteristic responses of an opioid receptor or receptor subtype. As anon-limiting example, an antagonist may competitively ornon-competitively bind to an opioid receptor, an agonist or partialagonist (or other ligand) of a receptor, and/or a downstream signalingmolecule to inhibit a receptor's function.

An inverse agonist able to block or inhibit a constitutive activity ofan opioid receptor may also be used. An inverse agonist maycompetitively or non-competitively bind to an opioid receptor and/or adownstream signaling molecule to inhibit a receptor's function.Non-limiting examples of inverse agonists for use in the disclosedmethods include ICI-174864 (N,N-diallyl-Tyr-Aib-Aib-Phe-Leu),RTI-5989-1, RTI-5989-23, and RTI-5989-25 (see Zaki et al. J. Pharmacol.Exp. Therap. 298(3): 1015-1020, 2001).

Additional embodiments of the disclosure include a combination of anangiotensin agent with an additional agent such as acetylcholine or areported modulator of an androgen receptor. Non-limiting examplesinclude the androgen receptor agonists dehydroepiandrosterone (DHEA) andDHEA sulfate (DHEAS).

Alternatively, the neurogenic agent in combination with an angiotensinagent may be an enzymatic inhibitor, such as a reported inhibitor of HMGCoA reductase. Non-limiting examples of such inhibitors includeatorvastatin (CAS RN 134523-00-5), cerivastatin (CAS RN 145599-86-6),crilvastatin (CAS RN 120551-59-9), fluvastatin (CAS RN 93957-54-1) andfluvastatin sodium (CAS RN 93957-55-2), simvastatin (CAS RN 79902-63-9),lovastatin (CAS RN 75330-75-5), pravastatin (CAS RN 81093-37-0) orpravastatin sodium, rosuvastatin (CAS RN 287714-41-4), and simvastatin(CAS RN 79902-63-9). Formulations containing one or more of suchinhibitors may also be used in a combination. Non-limiting examplesinclude formulations comprising lovastatin such as Advicor® (anextended-release, niacin containing formulation) or Altocor® (anextended release formulation); and formulations comprising simvastatinsuch as Vytorin® (combination of simvastatin and ezetimibe).

In other non-limiting embodiments, the neurogenic agent in combinationwith an angiotensin agent may be a reported Rho kinase inhibitor.Non-limiting examples of such an inhibitor include fasudil (CAS RN103745-39-7); fasudil hydrochloride (CAS RN 105628-07-7); the metaboliteof fasudil, which is hydroxyfasudil (see Shimokawa et al.“Rho-kinase-mediated pathway induces enhanced myosin light chainphosphorylations in a swine model of coronary artery spasm.” CardiovascRes. 1999 43:1029-1039), Y 27632 (CAS RN 138381-45-0); a fasudil analogthereof such as(S)-Hexahydro-1-(4-ethenylisoquinoline-5-sulfonyl)-2-methyl-1H-1,4-diazepine,(S)-hexahydro-4-glycyl-2-methyl-1-(4-methylisoquinoline-5-sulfonyl)-1H-1,4-diazepine,or (S)-(+)-2-methyl-1-[(4-methyl-5-isoquinoline)sulfonyl]-homopiperazine(also known as H-152P; see Sasaki et al. “The novel and specificRho-kinase inhibitor(S)-(+)-2-methyl-1-[(4-methyl-5-isoquinoline)sulfonyl]-homopiperazine asa probing molecule for Rho-kinase-involved pathway.” Pharmacol Ther.2002 93(2-3):225-32); or a substituted isoquinolinesulfonamide compoundas disclosed in U.S. Pat. No. 6,906,061.

Furthermore, the neurogenic agent in combination with an angiotensinagent may be a reported GSK-3 inhibitor or modulator. In somenon-limiting embodiments, the reported GSK3-beta modulator is apaullone, such as alsterpaullone, kenpaullone(9-bromo-7,12-dihydroindolo[3,2-d][1]benzazepin-6(5H)-one),gwennpaullone (see Knockaert et al. “Intracellular Targets of Paullones.Identification following affinity purification on immobilizedinhibitor.” J Biol Chem. 2002 277(28):25493-501), azakenpaullone (seeKunick et al. “1-Azakenpaullone is a selective inhibitor of glycogensynthase kinase-3 beta.” Bioorg Med Chem Lett. 2004 14(2):413-6), or thecompounds described in U.S. Publication No. 20030181439; InternationalPublication No. WO 01/60374; Leost et al., Eur. J. Biochem.267:5983-5994 (2000); Kunick et al., J Med Chem.; 47(1): 22-36 (2004);or Shultz et al., J. Med. Chem. 42:2909-2919 (1999); an anticonvulsant,such as lithium or a derivative thereof (e.g., a compound described inU.S. Pat. Nos. 1,873,732; 3,814,812; and 4,301,176); carbemazepine,valproic acid or a derivative thereof (e.g., valproate, or a compounddescribed in Werstuck et al., Bioorg Med Chem Lett., 14(22): 5465-7(2004)); lamotrigine; SL 76002 (Progabide), gabapentin; tiagabine; orvigabatrin; a maleimide or a related compound, such as Ro 31-8220,SB-216763, SB-410111, SB-495052, or SB-415286, or a compound described,e.g., in U.S. Pat. No. 6,719,520; U.S. Publication No. 20040010031;International Publication Nos. WO-2004072062; WO-03082859; WO-03104222;WO-03103663, WO-03095452, WO-2005000836; WO 0021927; WO-03076398;WO-00021927; WO-00038675; or WO-03076442; or Coghlan et al., Chemistry &Biology 7: 793 (2000); a pyridine or pyrimidine derivative, or a relatedcompound (such as 5-iodotubercidin, GI 179186X, GW 784752× and GW784775X, and compounds described, e.g., in U.S. Pat. Nos. 6,489,344;6,417,185; and 6153618; U.S. Publication Nos. 20050171094; and20030130289; European Patent Nos. EP-01454908, EP-01454910, EP-01295884,EP-01295885; and EP-01460076; EP-01454900; International PublicationNos. WO 01/70683; WO 01/70729; WO 01/70728; WO 01/70727; WO 01/70726; WO01/70725; WO-00218385; WO-00218386; WO-03072579; WO-03072580;WO-03027115; WO-03027116; WO-2004078760; WO-2005037800, WO-2004026881,WO-03076437, WO-03029223; WO-2004098607; WO-2005026155; WO-2005026159;WO-2005025567; WO-03070730; WO-03070729; WO-2005019218; WO-2005019219;WO-2004013140; WO-2004080977; WO-2004026229, WO-2004022561; WO-03080616;WO-03080609; WO-03051847; WO-2004009602; WO-2004009596; WO-2004009597;WO-03045949; WO-03068773; WO-03080617; WO 99/65897; WO 00/18758;WO0307073; WO-00220495; WO-2004043953, WO-2004056368, WO-2005012298,WO-2005012262, WO-2005042525, WO-2005005438, WO-2004009562, WO-03037877;WO-03037869; WO-03037891; WO-05012307; WO-05012304 and WO 98/16528; andin Massillon et al., Biochem J 299:123-8 (1994)); a pyrazine derivative,such as Aloisine A®(7-n-butyl-6-(4-hydroxyphenyl)[5H]pyrrolo[2,3-b]pyrazine) or a compounddescribed in International Publication Nos. WO-00144206; WO0144246; orWO-2005035532; a thiadiazole or thiazole, such as TDZD-8(benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione); OTDZT(4-dibenzyl-5-oxothiadiazolidine-3-thione); or a related compounddescribed, e.g., in U.S. Pat. Nos. 6,645,990 or 6,762,179; U.S.Publication No. 20010039275; International Publication Nos. WO 01/56567,WO-03011843, WO-03004478, or WO-03089419; or Mettey, Y., et al., J. Med.Chem. 46, 222 (2003); TWS119 or a related compound, such as a compounddescribed in Ding et al., Proc Natl Acad Sci USA., 100(13): 7632-7(2003); an indole derivative, such as a compound described inInternational Publication Nos. WO-03053330, WO-03053444, WO-03055877,WO-03055492, WO-03082853, or WO-2005027823; a pyrazine or pyrazolederivative, such as a compound described in U.S. Pat. Nos. 6,727,251,6,696,452, 6,664,247, 666,073, 6,656,939, 6,653,301, 6,653,300,6,638,926, 6,613,776, or 6,610,677; or International Publication Nos.WO-2005002552, WO-2005002576, or WO-2005012256; a compound described inU.S. Pat. Nos. 6,719,520; 6,498,176; 6,800,632; or 6,872,737; U.S.Publication Nos. 20050137201; 20050176713; 20050004125; 20040010031;20030105075; 20030008866; 20010044436; 20040138273; or 20040214928;International Publication Nos. WO 99/21859; WO-00210158; WO-05051919;WO-00232896; WO-2004046117; WO-2004106343; WO-00210141; WO-00218346; WO00/21927; WO 01/81345; WO 01/74771; WO 05/028475; WO 01/09106; WO00/21927; WO01/41768; WO 00/17184; WO 04/037791; WO-04065370; WO01/37819; WO 01/42224; WO 01/85685; WO 04/072063; WO-2004085439;WO-2005000303; WO-2005000304; or WO 99/47522; or Naerum, L., et al.,Bioorg. Med. Chem. Lett. 12, 1525 (2002); CP-79049, GI 179186X, GW784752X, GW 784775X, AZD-1080, AR-014418, SN-8914, SN-3728, OTDZT,Aloisine A, TWS119, CHIR98023, CHIR99021, CHIR98014, CHIR98023,5-iodotubercidin, Ro 31-8220, SB-216763, SB-410111, SB-495052,SB-415286, alsterpaullone, kenpaullone, gwennpaullone, LY294002,wortmannin, sildenafil, CT98014, CT-99025, flavoperidol, or L803-mts.

In yet further embodiments, the neurogenic agent used in combinationwith an angiotensin agent may be a reported glutamate modulator ormetabotropic glutamate (mGlu) receptor modulator. In some embodiments,the reported mGlu receptor modulator is a Group II modulator, havingactivity against one or more Group II receptors (mGlu₂ and/or mGlu₃).Embodiments include those where the Group II modulator is a Group IIagonist. Non-limiting examples of Group II agonists include: (i)(1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD), a broadspectrum mGlu agonist having substantial activity at Group I and IIreceptors; (ii) (−)-2-thia-4-aminobicyclo-hexane-4,6-dicarboxylate(LY389795), which is described in Monn et al., J. Med. Chem.,42(6):1027-40 (1999); (iii) compounds described in US App. No.20040102521 and Pellicciari et al., J. Med. Chem., 39, 2259-2269 (1996);and (iv) the Group II-specific modulators described below.

Non-limiting examples of reported Group II antagonists include: (i)phenylglycine analogues, such as(RS)-alpha-methyl-4-sulphonophenylglycine (MSPG),(RS)-alpha-methyl-4-phosphonophenylglycine (MPPG), and(RS)-alpha-methyl-4-tetrazolylphenylglycine (MTPG), described in Jane etal., Neuropharmacology 34: 851-856 (1995); (ii) LY366457, which isdescribed in O'Neill et al., Neuropharmacol., 45(5): 565-74 (2003);(iii) compounds described in US App Nos. 20050049243, 20050119345 and20030157647; and (iv) the Group II-specific modulators described below.

In some non-limiting embodiments, the reported Group II modulator is aGroup II-selective modulator, capable of modulating mGlu₂ and/or mGlu₃under conditions where it is substantially inactive at other mGlusubtypes (of Groups I and III). Examples of Group II-selectivemodulators include compounds described in Monn, et al., J. Med. Chem.,40, 528-537 (1997); Schoepp, et al., Neuropharmacol., 36, 1-11 (1997)(e.g., 1S,2S,5R,6S-2-aminobicyclohexane-2,6-dicarboxylate); and Schoepp,Neurochem. Int., 24, 439 (1994).

Non-limiting examples of reported Group II-selective agonists include(i) (+)-2-aminobicyclohexane-2,6-dicarboxylic acid (LY354740), which isdescribed in Johnson et al., Drug Metab. Disposition, 30(1): 27-33(2002) and Bond et al., NeuroReport 8: 1463-1466 (1997), and issystemically active after oral administration (e.g., Grillon et al.,Psychopharmacol. (Berl), 168: 446-454 (2003)); (ii)(−)-2-oxa-4-aminobicyclohexane-4,6-dicarboxylic acid (LY379268), whichis described in Monn et al., J. Med. Chem. 42: 1027-1040 (1999) and U.S.Pat. No. 5,688,826. LY379268 is readily permeable across the blood-brainbarrier, and has EC₅₀ values in the low nanomolar range (e.g., belowabout 10 nM, or below about 5 nM) against human mGlu₂ and mGlu₃receptors in vitro; (iii) (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate((2R,4R)-APDC), which is described in Monn et al., J. Med. Chem. 39:2990 (1996) and Schoepp et al., Neuropharmacology, 38: 1431 (1999); (iv)(1S,3S)-1-aminocyclopentane-1,3-dicarboxylic acid ((1S,3S)-ACPD),described in Schoepp, Neurochem. Int., 24: 439 (1994); (v)(2R,4R)-4-aminopyrrolidine-2,4-dicarboxylic acid ((2R,4R)-APDC),described in Howson and Jane, British Journal of Pharmacology, 139,147-155 (2003); (vi) (2S,1′S,2′S)-2-(carboxycyclopropyl)-glycine(L-CCG-I), described in Brabet et al., Neuropharmacology 37: 1043-1051(1998); (vii) (2S,2′R,3′R)-2-(2′,3′-dicarboxycyclopropyl)glycine(DCG-IV), described in Hayashi et al., Nature, 366, 687-690 (1993);(viii) 1S,2S,5R,6S-2-aminobicyclohexane-2,6-dicarboxylate, described inMonn, et al., J. Med. Chem., 40, 528 (1997) and Schoepp, et al.,Neuropharmacol., 36, 1 (1997); and (ix) compounds described in US App.No. 20040002478; U.S. Pat. Nos. 6,204,292, 6,333,428, 5,750,566 and6,498,180; and Bond et al., Neuroreport 8: 1463-1466 (1997).

Non-limiting examples of reported Group II-selective antagonists usefulin methods provided herein include the competitive antagonist(2S)-2-amino-2-(1S,2S-2-carboxycycloprop-1-yl)-3-(xanth-9-yl)propanoicacid (LY341495), which is described, e.g., in Kingston et al.,Neuropharmacology 37: 1-12 (1998) and Monn et al., J Med Chem 42:1027-1040 (1999). LY341495 is readily permeably across the blood-brainbarrier, and has IC₅₀ values in the low nanomolar range (e.g., belowabout 10 nM, or below about 5 nM) against cloned human mGlu₂ and mGlu₃receptors. LY341495 has a high degree of selectivity for Group IIreceptors relative to Group I and Group III receptors at lowconcentrations (e.g., nanomolar range), whereas at higher concentrations(e.g., above 1 μM), LY341495 also has antagonist activity against mGlu₇and mGlu₈, in addition to mGlu_(2/3). LY341495 is substantially inactiveagainst KA, AMPA, and NMDA iGlu receptors.

Additional non-limiting examples of reported Group II-selectiveantagonists include the following compounds, indicated by chemical nameand/or described in the cited references: (i)α-methyl-L-(carboxycyclopropyl)glycine (CCG); (ii)(2S,3S,4S)-2-methyl-2-(carboxycyclopropyl)glycine (MCCG); (iii)(1R,2R,3R,5R,6R)-2-amino-3-(3,4-dichlorobenzyloxy)-6fluorobicyclohexane-2,6-dicarboxylic acid (MGS0039), which is describedin Nakazato et al., J. Med. Chem., 47(18):4570-87 (2004); (iv) ann-hexyl, n-heptyl, n-octyl, 5-methylbutyl, or 6-methylpentyl esterprodrug of MGS0039; (v) MGS0210(3-(3,4-dichlorobenzyloxy)-2-amino-6-fluorobicyclohexane-2,6-dicarboxylicacid n-heptyl ester); (vi) (RS)-1-amino-5-phosphonoindan-1-carboxylicacid (APICA), which is described in Ma et al., Bioorg. Med. Chem. Lett.,7: 1195 (1997); (vii) (2S)-ethylglutamic acid (EGLU), which is describedin Thomas et al., Br. J. Pharmacol. 117: 70P (1996); (viii)(2S,1′S,2′S,3′R)-2-(2′-carboxy-3′-phenylcyclopropyl)glycine (PCCG-IV);and (ix) compounds described in U.S. Pat. No. 6,107,342 and US App No.20040006114. APICA has an IC₅₀ value of approximately 30 μM againstmGluR₂ and mGluR₃, with no appreciable activity against Group I or GroupIII receptors at sub-mM concentrations.

In some non-limiting embodiments, a reported Group II-selectivemodulator is a subtype-selective modulator, capable of modulating theactivity of mGlu₂ under conditions in which it is substantially inactiveat mGlu₃ (mGlu₂-selective), or vice versa (mGlu₃-selective).Non-limiting examples of subtype-selective modulators include compoundsdescribed in U.S. Pat. No. 6,376,532 (mGlu₂-selective agonists) and USApp No. 20040002478 (mGlu₃-selective agonists). Additional non-limitingexamples of subtype-selective modulators include allosteric mGlureceptor modulators (mGlu₂ and mGlu₃) and NAAG-related compounds(mGlu₃), such as those described below.

In other non-limiting embodiments, a reported Group II modulator is acompound with activity at Group I and/or Group III receptors, inaddition to Group II receptors, while having selectivity with respect toone or more mGlu receptor subtypes. Non-limiting examples of suchcompounds include: (i) (2S,3S,4S)-2-(carboxycyclopropyl)glycine(L-CCG-1) (Group I/Group II agonist), which is described in Nicoletti etal., Trends Neurosci. 19: 267-271 (1996), Nakagawa, et al., Eur. J.Pharmacol., 184, 205 (1990), Hayashi, et al., Br. J. Pharmacol., 107,539 (1992), and Schoepp et al., J. Neurochem., 63., page 769-772 (1994);(ii) (S)-4-carboxy-3-hydroxyphenylglycine (4C₃HPG) (Group IIagonist/Group I competitive antagonist); (iii) gamma-carboxy-L-glutamicacid (GLA) (Group II antagonist/Group III partial agonist/antagonist);(iv) (2S,2′R,3′R)-2-(2,3-dicarboxycyclopropyl)glycine (DCG-IV) (Group IIagonist/Group III antagonist), which is described in Ohfune et al,Bioorg. Med. Chem. Lett., 3: 15 (1993); (v)(RS)-a-methyl-4-carboxyphenylglycine (MCPG) (Group I/Group IIcompetitive antagonist), which is described in Eaton et al., Eur. J.Pharmacol., 244: 195 (1993), Collingridge and Watkins, TiPS, 15: 333(1994), and Joly et al., J. Neurosci., 15: 3970 (1995); and (vi) theGroup II/III modulators described in U.S. Pat. Nos. 5,916,920,5,688,826, 5,945,417, 5,958,960, 6,143,783, 6,268,507, 6,284,785.

In some non-limiting embodiments, the reported mGlu receptor modulatorcomprises (S)-MCPG (the active isomer of the Group I/Group IIcompetitive antagonist (RS)-MCPG) substantially free from (R)-MCPG.(S)-MCPG is described, e.g., in Sekiyama et al., Br. J. Pharmacol., 117:1493 (1996) and Collingridge and Watkins, TiPS, 15: 333 (1994).

Additional non-limiting examples of reported mGlu modulators useful inmethods disclosed herein include compounds described in U.S. Pat. Nos.6,956,049, 6,825,211, 5,473,077, 5,912,248, 6,054,448, and 5,500,420; USApp Nos. 20040077599, 20040147482, 20040102521, 20030199533 and20050234048; and Intl Pub/App Nos. WO 97/19049, WO 98/00391, andEP0870760.

In some non-limiting embodiments, the reported mGlu receptor modulatoris a prodrug, metabolite, or other derivative ofN-acetylaspartylglutamate (NAAG), a peptide neurotransmitter in themammalian CNS that is a highly selective agonist for mGluR₃ receptors,as described in Wroblewska et al., J. Neurochem., 69(1): 174-181 (1997).In other embodiments, the mGlu modulator is a compound that modulatesthe levels of endogenous NAAG, such as an inhibitor of the enzymeN-acetylated-alpha-linked-acidic dipeptidase (NAALADase), whichcatalyzes the hydrolysis of NAAG to N-acetyl-aspartate and glutamate.Examples of NAALADase inhibitors include 2-PMPA(2-(phosphonomethyl)pentanedioic acid), which is described in Slusher etal., Nat. Med., 5(12): 1396-402 (1999); and compounds described in J.Med. Chem. 39: 619 (1996), US Pub. No. 20040002478, and U.S. Pat. Nos.6,313,159, 6,479,470, and 6,528,499. In some embodiments, the mGlumodulator is the mGlu₃-selective antagonist, beta-NAAG.

Additional non-limiting examples of reported glutamate modulatorsinclude memantine (CAS RN 19982-08-2), memantine hydrochloride (CAS RN41100-52-1), and riluzole (CAS RN 1744-22-5).

In some non-limiting embodiments, a reported Group II modulator isadministered in combination with one or more additional compoundsreported as active against a Group I and/or a Group III mGlu receptor.For example, in some cases, methods comprise modulating the activity ofat least one Group I receptor and at least one Group II mGlu receptor(e.g., with a compound described herein). Examples of compounds usefulin modulating the activity of Group I receptors include GroupI-selective agonists, such as (i) trans-azetidine-2,4-dicarboxylic acid(tADA), which is described in Kozikowski et al., J. Med. Chem., 36: 2706(1993) and Manahan-Vaughan et al., Neuroscience, 72: 999 (1996); (ii)(RS)-3,5-dihydroxyphenylglycine (DHPG), which is described in Ito etal., NeuroReport 3: 1013 (1992); or a composition comprising (S)-DHPGsubstantially free of (R)-DHPG, as described, e.g., in Baker et al.,Bioorg. Med. Chem. Lett. 5: 223 (1995); (iii)(RS)-3-hydroxyphenylglycine, which is described in Birse et al.,Neuroscience 52: 481 (1993); or a composition comprising(S)-3-hydroxyphenylglycine substantially free of(R)-3-hydroxyphenylglycine, as described, e.g., in Hayashi et al., J.Neurosci., 14: 3370 (1994); (iv) and (S)-homoquisqualate, which isdescribed in Porter et al., Br. J. Pharmacol., 106: 509 (1992).

Additional non-limiting examples of reported Group I modulators include(i) Group I agonists, such as (RS)-3,5-dihydroxyphenylglycine, describedin Brabet et al., Neuropharmacology, 34, 895-903, 1995; and compoundsdescribed in U.S. Pat. Nos. 6,399,641 and 6,589,978, and US Pub No.20030212066; (ii) Group I antagonists, such as(S)-4-carboxy-3-hydroxyphenylglycine;7-(hydroxyimino)cyclopropa-β-chromen-1α-carboxylate ethyl ester;(RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA);2-methyl-6(phenylethynyl)pyridine (MPEP);2-methyl-6-(2-phenylethenyl)pyridine (SIB-1893);6-methyl-2-(phenylazo)-3-pyridinol (SIB-1757);(Sα-amino-4-carboxy-2-methylbenzeneacetic acid; and compounds describedin U.S. Pat. Nos. 6,586,422, 5,783,575, 5,843,988, 5,536,721, 6,429,207,5,696,148, and 6,218,385, and US Pub Nos. 20030109504, 20030013715,20050154027, 20050004130, 20050209273, 20050197361, and 20040082592;(iii) mGlu₅-selective agonists, such as(RS)-2-chloro-5-hydroxyphenylglycine (CHPG); and (iv) mGlu₅-selectiveantagonists, such as 2-methyl-6-(phenylethynyl)-pyridine (MPEP); andcompounds described in U.S. Pat. No. 6,660,753; and US Pub Nos.20030195139, 20040229917, 20050153986, 20050085514, 20050065340,20050026963, 20050020585, and 20040259917.

Non-limiting examples of compounds reported to modulate Group IIIreceptors include (i) the Group III-selective agonists(L)-2-amino-4-phosphonobutyric acid (L-AP4), described in Knopfel etal., J. Med Chem., 38, 1417-1426 (1995); and(S)-2-amino-2-methyl-4-phosphonobutanoic acid; (ii) the GroupIII-selective antagonists (RS)-α-cyclopropyl-4-phosphonophenylglycine;(RS)-α-methylserine-O-phosphate (MSOP); and compounds described in USApp. No. 20030109504; and (iii)(1S,3R,4S)-1-aminocyclopentane-1,2,4-tricarboxylic acid (ACPT-I).

In additional embodiments, the neurogenic agent used in combination withan angiotensin agent may be a reportedalpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)modulator. Non-limiting examples include CX-516 or ampalex (CAS RN154235-83-3), Org-24448 (CAS RN 211735-76-1), LY451395(2-propanesulfonamide,N-[(2R)-2-[4′-[2-[methylsulfonyl)amino]ethyl][1,1′-biphenyl]-4-yl]propyl]-),LY-450108 (see Jhee et al. “Multiple-dose plasma pharmacokinetic andsafety study of LY450108 and LY451395 (AMPA receptor potentiators) andtheir concentration in cerebrospinal fluid in healthy human subjects.” JClin Pharmacol. 2006 46(4):424-32), and CX717. Additional examples ofreported antagonists include irampanel (CAS RN 206260-33-5) and E-2007.

Further non-limiting examples of reported AMPA receptor antagonists foruse in combinations include YM90K (CAS RN 154164-30-4), YM872 orzonampanel (CAS RN 210245-80-0), NBQX (or2,3-dioxo-6-nitro-7-sulfamoylbenzo[f]quinoxaline; CAS RN 118876-58-7),PNQX(1,4,7,8,9,10-hexahydro-9-methyl-6-nitropyrido[3,4-f]quinoxaline-2,3-dione),and ZK200775([1,2,3,4-tetrahydro-7-morpholinyl-2,3-dioxo-6-(fluoromethyl)quinoxalin-1-yl]methylphosphonate).

In additional embodiments, a neurogenic agent used in combination withan angiotensin agent may be a reported muscarinic agent. Non-limitingexamples of a reported muscarinic agent include a muscarinic agonistsuch as milameline (CI-979), or a structurally or functionally relatedcompound disclosed in U.S. Pat. Nos. 4,786,648, 5,362,860, 5,424,301,5,650,174, 4,710,508, 5,314,901, 5,356,914, or 5,356,912; or xanomeline,or a structurally or functionally related compound disclosed in U.S.Pat. Nos. 5,041,455, 5,043,345, or 5,260,314.

Other non-limiting examples include a muscarinic agent such asalvameline (LU 25-109), or a functionally or structurally compounddisclosed in U.S. Pat. Nos. 6,297,262, 4,866,077, RE36,374, 4,925,858,PCT Publication No. WO 97/17074, or in Moltzen et al., J Med Chem. 1994Nov. 25; 37(24):4085-99;2,8-dimethyl-3-methylene-1-oxa-8-azaspiro[4.5]decane (YM-796) or YM-954,or a functionally or structurally related compound disclosed in U.S.Pat. No. 4,940,795, RE34,653, 4,996,210, 5,041,549, 5,403,931, or5,412,096, or in Wanibuchi et al., Eur. J. Pharmacol., 187, 479-486(1990); cevimeline (AF102B), or a functionally or structurally compounddisclosed in U.S. Pat. Nos. 4,855,290, 5,340,821, 5,580,880 (AmericanHome Products), or 4,981,858 (optical isomers of AF102B); sabcomeline(SB 202026), or a functionally or structurally related compounddescribed in U.S. Pat. Nos. 5,278,170, RE35,593, 6,468,560, 5,773,619,5,808,075, 5,545,740, 5,534,522, or 6,596,869, U.S. Patent PublicationNos. 2002/0127271, 2003/0129246, 2002/0150618, 2001/0018074,2003/0157169, or 2001/0003588, Bromidge et al., J Med Chem. 19;40(26):4265-80 (1997), or Harries et al., British J. Pharm., 124,409-415 (1998); talsaclidine (WAL 2014 FU), or a functionally orstructurally compound disclosed in U.S. Pat. Nos. 5,451,587, 5,286,864,5,508,405, 5,451,587, 5,286,864, 5,508,405, or 5,137,895, or inPharmacol. Toxicol., 78, 59-68 (1996); or a1-methyl-1,2,5,6-tetrahydropyridyl-1,2,5-thiadiazole derivative, such astetra(ethyleneglycol)(4-methoxy-1,2,5-thiadiazol-3-yl)[3-(1-methyl-1,2,5,6-tetrahydropyrid-3-yl)-1,2,5-thiadiazol-4-yl]ether,or a compound that is functionally or structurally related to a1-methyl-1,2,5,6-tetrahydropyridyl-1,2,5-thiadiazole derivative asprovided by Cao et al. (“Synthesis and biological characterization of1-methyl-1,2,5,6-tetrahydropyridyl-1,2,5-thiadiazole derivatives asmuscarinic agonists for the treatment of neurological disorders.” J.Med. Chem. 46(20):4273-4286, 2003).

Yet additional non-limiting examples include besipiridine, SR-46559,L-689,660, S-9977-2, AF-102, thiopilocarpine, or an analog of clozapine,such as a pharmaceutically acceptable salt, ester, amide, or prodrugform thereof, or a diaryl[a,d]cycloheptene, such as an amino substitutedform thereof, or N-desmethylclozapine, which has been reported to be ametabolite of clozapine, or an analog or related compound disclosed inUS 2005/0192268 or WO 05/63254.

In other embodiments, the muscarinic agent is an m₁ receptor agonistselected from 55-LH-3B, 55-LH-25A, 55-LH-30B, 55-LH-4-1A,40-LH-67,55-LH-15A, 55-LH-16B, 55-LH-11C, 55-LH-31A, 55-LH-46, 55-LH-47,55-LH-4-3A, or a compound that is functionally or structurally relatedto one or more of these agonists disclosed in US 2005/0130961 orWO04/087158.

In additional embodiments, the muscarinic agent is a benzimidazolidinonederivative, or a functionally or structurally compound disclosed in U.S.Pat. No. 6,951,849, US 2003/0100545, WO 04/089942, or WO 03/028650; aspiroazacyclic compound, or a functionally or structurally relatedcompound like 1-oxa-3,8-diaza-spiro[4,5]decan-2-one or a compounddisclosed in U.S. Pat. No. 6,911,452 or WO 03/057698; or atetrahydroquinoline analog, or a functionally or structurally compounddisclosed in US 2003/0176418, US 2005/0209226, or WO 03/057672.

In yet additional embodiments, the neurogenic agent in combination withan angiotensin agent is a reported HDAC inhibitor. The term “HDAC”refers to any one of a family of enzymes that remove acetyl groups fromthe epsilon-amino groups of lysine residues at the N-terminus of ahistone. An HDAC inhibitor refers to compounds capable of inhibiting,reducing, or otherwise modulating the deacetylation of histones mediatedby a histone deacetylase. Non-limiting examples of a reported HDACinhibitor include a short-chain fatty acid, such as butyric acid,phenylbutyrate (PB), 4-phenylbutyrate (4-PBA), pivaloyloxymethylbutyrate (Pivanex, AN-9), isovalerate, valerate, valproate, valproicacid, propionate, butyramide, isobutyramide, phenylacetate,3-bromopropionate, or tributyrin; a compound bearing a hydroxyamic acidgroup, such as suberoylanlide hydroxamic acid (SAHA), trichostatin A(TSA), trichostatin C (TSC), salicylhydroxamic acid, oxamflatin, subericbishydroxamic acid (SBHA), m-carboxy-cinnamic acid bishydroxamic acid(CBHA), pyroxamide (CAS RN 382180-17-8), diethylbis-(pentamethylene-N,N-dimethylcarboxamide) malonate (EMBA), azelaicbishydroxamic acid (ABHA), azelaic-1-hydroxamate-9-anilide (AAHA),6-(3-chlorophenylureido) carpoic hydroxamic acid, or A-161906; a cyclictetrapeptide, such as depsipeptide (FK228), FR225497, trapoxin A,apicidin, chlamydocin, or HC-toxin; a benzamide, such as MS-275;depudecin, a sulfonamide anilide (e.g., diallyl sulfide), BL1521,curcumin (diferuloylmethane), CI-994 (N-acetyldinaline), spiruchostatinA, scriptaid, carbamazepine (CBZ), or a related compound; a compoundcomprising a cyclic tetrapeptide group and a hydroxamic acid group(examples of such compounds are described in U.S. Pat. Nos. 6,833,384and 6,552,065); a compound comprising a benzamide group and a hydroxamicacid group (examples of such compounds are described in Ryu et al.,Cancer Lett. 2005 Jul. 9 (epub), Plumb et al., Mol Cancer Ther.,2(8):721-8 (2003), Ragno et al., J Med Chem., 47(6):1351-9 (2004), Maiet al., J Med Chem., 47(5):1098-109 (2004), Mai et al., J Med Chem.,46(4):512-24 (2003), Mai et al., J Med Chem., 45(9):1778-84 (2002),Massa et al., J Med Chem., 44(13):2069-72 (2001), Mai et al., J MedChem., 48(9):3344-53 (2005), and Mai et al., J Med Chem., 46(23):4826-9(2003)); a compound described in U.S. Pat. Nos. 6,897,220, 6,888,027,5,369,108, 6,541,661, 6,720,445, 6,562,995, 6,777,217, or 6,387,673, orU.S. Patent Publication Nos. 20050171347, 20050165016, 20050159470,20050143385, 20050137234, 20050137232, 20050119250, 20050113373,20050107445, 20050107384, 20050096468, 20050085515, 20050032831,20050014839, 20040266769, 20040254220, 20040229889, 20040198830,20040142953, 20040106599, 20040092598, 20040077726, 20040077698,20040053960, 20030187027, 20020177594, 20020161045, 20020119996,20020115826, 20020103192, or 20020065282; FK228, AN-9, MS-275, CI-994,SAHA, G2M-777, PXD-101, LBH-589, MGCD-0103, MK0683, sodiumphenylbutyrate, CRA-024781, and derivatives, salts, metabolites,prodrugs, and stereoisomers thereof; and a molecule that inhibits thetranscription and/or translation of one or more HDACs.

Additional non-limiting examples include a reported HDac inhibitorselected from ONO-2506 or arundic acid (CAS RN 185517-21-9); MGCD0103(see Gelmon et al. “Phase I trials of the oral histone deacetylase(HDAC) inhibitor MGCD0103 given either daily or 3× weekly for 14 daysevery 3 weeks in patients (pts) with advanced solid tumors.” Journal ofClinical Oncology, 2005 ASCO Annual Meeting Proceedings. 23(16S, June 1Supplement), 2005: 3147 and Kalita et al. “Pharmacodynamic effect ofMGCD0103, an oral isotype-selective histone deacetylase (HDAC)inhibitor, on HDAC enzyme inhibition and histone acetylation inductionin Phase I clinical trials in patients (pts) with advanced solid tumorsor non-Hodgkin's lymphoma (NHL)” Journal of Clinical Oncology, 2005 ASCOAnnual Meeting Proceedings. 23(16S, Part I of II, June 1 Supplement),2005: 9631), a reported thiophenyl derivative of benzamide HDacinhibitor as presented at the 97th American Association for CancerResearch (AACR) Annual Meeting in Washington, D.C. in a poster titled“Enhanced Isotype-Selectivity and Antiproliferative Activity ofThiophenyl Derivatives of Benzamide HDAC Inhibitors In Human CancerCells,” (abstract #4725), and a reported HDac inhibitor as described inU.S. Pat. No. 6,541,661; SAHA or vorinostat (CAS RN 149647-78-9); PXD101or PXD 101 or PX 105684 (CAS RN 414864-00-9), CI-994 or tacedinaline(CAS RN 112522-64-2), MS-275 (CAS RN 209783-80-2), or an inhibitorreported in WO 2005/108367.

In other embodiments, the neurogenic agent in combination with anangiotensin agent is a reported GABA modulator which modulates GABAreceptor activity at the receptor level (e.g., by binding directly toGABA receptors), at the transcriptional and/or translational level(e.g., by preventing GABA receptor gene expression), and/or by othermodes (e.g., by binding to a ligand or effector of a GABA receptor, orby modulating the activity of an agent that directly or indirectlymodulates GABA receptor activity). Non-limiting examples of GABA-Areceptor modulators useful in methods described herein includetriazolophthalazine derivatives, such as those disclosed in WO 99/25353,and WO/98/04560; tricyclic pyrazolo-pyridazinone analogues, such asthose disclosed in WO 99/00391; fenamates, such as those disclosed inU.S. Pat. No. 5,637,617; triazolo-pyridazine derivatives, such as thosedisclosed in WO 99/37649, WO 99/37648, and WO 99/37644;pyrazolo-pyridine derivatives, such as those disclosed in WO 99/48892;nicotinic derivatives, such as those disclosed in WO 99/43661 and U.S.Pat. No. 5,723,462; muscimol, thiomuscimol, and compounds disclosed inU.S. Pat. No. 3,242,190; baclofen and compounds disclosed in U.S. Pat.No. 3,471,548; phaclofen; quisqualamine; ZAPA; zaleplon; THIP;imidazole-4-acetic acid (IMA); (+)-bicuculline; gabalinoleamide;isoguvicaine; 3-aminopropane sulphonic acid; piperidine-4-sulphonicacid; 4,5,6,7-tetrahydro-[5,4-c]-pyridin-3-ol; SR 95531; RU5315; CGP55845; CGP 35348; FG 8094; SCH 50911; NG2-73; NGD-96-3; pricrotoxin andother bicyclophosphates disclosed in Bowery et al., Br. J. Pharmacol.,57; 435 (1976).

Additional non-limiting examples of GABA-A modulators include compoundsdescribed in U.S. Pat. Nos. 6,503,925; 6,218,547; 6,399,604; 6,646,124;6,515,140; 6,451,809; 6,448,259; 6,448,246; 6,423,711; 6,414,147;6,399,604; 6,380,209; 6,353,109; 6,297,256; 6,297,252; 6,268,496;6,211,365; 6,166,203; 6,177,569; 6,194,427; 6,156,898; 6,143,760;6,127,395; 6,103,903; 6,103,731; 6,723,735; 6,479,506; 6,476,030;6,337,331; 6,730,676; 6,730,681; 6,828,322; 6,872,720; 6,699,859;6,696,444; 6,617,326; 6,608,062; 6,579,875; 6,541,484; 6,500,828;6,355,798; 6,333,336; 6,319,924; 6,303,605; 6,303,597; 6,291,460;6,255,305; 6,133,255; 6,872,731; 6,900,215; 6,642,229; 6,593,325;6,914,060; 6,914,063; 6,914,065; 6,936,608; 6,534,505; 6,426,343;6,313,125; 6,310,203; 6,200,975; 6,071,909; 5,922,724; 6,096,887;6,080,873; 6,013,799; 5,936,095; 5,925,770; 5,910,590; 5,908,932;5,849,927; 5,840,888; 5,817,813; 5,804,686; 5,792,766; 5,750,702;5,744,603; 5,744,602; 5,723,462; 5,696,260; 5,693,801; 5,677,309;5,668,283; 5,637,725; 5,637,724; 5,625,063; 5,610,299; 5,608,079;5,606,059; 5,604,235; 5,585,490; 5,510,480; 5,484,944; 5,473,073;5,463,054; 5,451,585; 5,426,186; 5,367,077; 5,328,912 5,326,868;5,312,822; 5,306,819; 5,286,860; 5,266,698; 5,243,049; 5,216,159;5,212,310; 5,185,446; 5,185,446; 5,182,290; 5,130,430; 5,095,015;20050014939; 20040171633; 20050165048; 20050165023; 20040259818; and20040192692.

In some embodiments, the GABA-A modulator is a subunit-selectivemodulator. Non-limiting examples of GABA-A modulator having specificityfor the alpha1 subunit include alpidem and zolpidem. Non-limitingexamples of GABA-A modulator having specificity for the alpha2 and/oralpha3 subunits include compounds described in U.S. Pat. Nos. 6,730,681;6,828,322; 6,872,720; 6,699,859; 6,696,444; 6,617,326; 6,608,062;6,579,875; 6,541,484; 6,500,828; 6,355,798; 6,333,336; 6,319,924;6,303,605; 6,303,597; 6,291,460; 6,255,305; 6,133,255; 6,900,215;6,642,229; 6,593,325; and 6,914,063. Non-limiting examples of GABA-Amodulator having specificity for the alpha2, alpha3 and/or alpha5subunits include compounds described in U.S. Pat. Nos. 6,730,676 and6,936,608. Non-limiting examples of GABA-A modulators having specificityfor the alpha5 subunit include compounds described in U.S. Pat. Nos.6,534,505; 6,426,343; 6,313,125; 6,310,203; 6,200,975 and 6,399,604.Additional non-limiting subunit selective GABA-A modulators includeCL218,872 and related compounds disclosed in Squires et al., Pharmacol.Biochem. Behav., 10: 825 (1979); and beta-carboline-3-carboxylic acidesters described in Nielsen et al., Nature, 286: 606 (1980).

In some embodiments, the GABA-A receptor modulator is a reportedallosteric modulator. In various embodiments, allosteric modulatorsmodulate one or more aspects of the activity of GABA at the target GABAreceptor, such as potency, maximal effect, affinity, and/orresponsiveness to other GABA modulators. In some embodiments, allostericmodulators potentiate the effect of GABA (e.g., positive allostericmodulators), and/or reduce the effect of GABA (e.g., inverse agonists).Non-limiting examples of benzodiazepine GABA-A modulators includealprazolam, bentazepam, bretazenil, bromazepam, brotizolam, cannazepam,chlordiazepoxide, clobazam, clonazepam, cinolazepam, clotiazepam,cloxazolam, clozapin, delorazepam, diazepam, dibenzepin, dipotassiumchlorazepat, divaplon, estazolam, ethyl-loflazepat, etizolam,fludiazepam, flumazenil, flunitrazepam, flurazepam 1HCl, flutoprazepam,halazepam, haloxazolam, imidazenil, ketazolam, lorazepam, loprazolam,lormetazepam, medazepam, metaclazepam, mexozolam, midazolam-HCl,nabanezil, nimetazepam, nitrazepam, nordazepam, oxazepam-tazepam,oxazolam, pinazepam, prazepam, quazepam, sarmazenil, suriclone,temazepam, tetrazepam, tofisopam, triazolam, zaleplon, zolezepam,zolpidem, zopiclone, and zopielon.

Additional non-limiting examples of benzodiazepine GABA-A modulatorsinclude Ro15-4513, CL218872, CGS 8216, CGS 9895, PK 9084, U-93631,beta-CCM, beta-CCB, beta-CCP, Ro 19-8022, CGS 20625, NNC 14-0590, Ru33-203, 5-amino-1-bromouracil, GYKI-52322, FG 8205, Ro 19-4603, ZG-63,RWJ46771, SX-3228, and L-655,078; NNC 14-0578, NNC 14-8198, andadditional compounds described in Wong et al., Eur J Pharmacol 209:319-325 (1995); Y-23684 and additional compounds in Yasumatsu et al., BrJ Pharmacol 111: 1170-1178 (1994); and compounds described in U.S. Pat.No. 4,513,135.

Non-limiting examples of barbiturate or barbituric acid derivativeGABA-A modulators include phenobarbital, pentobarbital, pentobarbitone,primidone, barbexaclon, dipropyl barbituric acid, eunarcon,hexobarbital, mephobarbital, methohexital, Na-methohexital,2,4,6(1H,3H,5)-pyrimidintrion, secbutabarbital and/or thiopental.

Non-limiting examples of neurosteroid GABA-A modulators includealphaxalone, allotetrahydrodeoxycorticosterone,tetrahydrodeoxycorticosterone, estrogen, progesterone3-beta-hydroxyandrost-5-en-17-on-3-sulfate, dehydroepianrosterone,eltanolone, ethinylestradiol, 5-pregnen-3-beta-ol-20 on-sulfate,5a-pregnan-3a-ol-20-one (5PG), allopregnanolone, pregnanolone, andsteroid derivatives and metabolites described in U.S. Pat. Nos.5,939,545, 5,925,630, 6,277,838, 6,143,736, RE35,517, 5,925,630,5,591,733, 5,232,917, 20050176976, WO 96116076, WO 98/05337, WO95/21617, WO 94/27608, WO 93/18053, WO 93/05786, WO 93/03732, WO91116897, EP01038880, and Han et al., J. Med. Chem., 36, 3956-3967(1993), Anderson et al., J. Med. Chem., 40, 1668-1681 (1997), Hogenkampet al., J. Med. Chem., 40, 61-72 (1997), Upasani et al., J. Med. Chem.,40, 73-84 (1997), Majewska et al., Science 232:1004-1007 (1986),Harrison et al., J. Pharmacol. Exp. Ther. 241:346-353 (1987), Gee etal., Eur. J. Pharmacol., 136:419-423 (1987) and Birtran et al., BrainRes., 561, 157-161 (1991).

Non-limiting examples of beta-carboline GABA-A modulators includeabecarnil, 3,4-dihydro-beta-carboline, gedocarnil,1-methyl-1-vinyl-2,3,4-trihydro-beta-carboline-3-carboxylic acid,6-methoxy-1,2,3,4-tetrahydro-beta-carboline,N—BOC-L-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid, tryptoline,pinoline, methoxyharmalan, tetrahydro-beta-carboline (THBC),1-methyl-THBC, 6-methoxy-THBC, 6-hydroxy-THBC, 6-methoxyharmalan,norharman, 3,4-dihydro-beta-carboline, and compounds described inNielsen et al., Nature, 286: 606 (1980).

In some embodiments, the GABA modulator modulates GABA-B receptoractivity. Non-limiting examples of reported GABA-B receptor modulatorsuseful in methods described herein include CGP36742; CGP-64213; CGP56999A; CGP 54433A; CGP 36742; SCH 50911; CGP 7930; CGP 13501; baclofenand compounds disclosed in U.S. Pat. No. 3,471,548; saclofen; phaclofen;2-hydroxysaclofen; SKF 97541; CGP 35348 and related compounds describedin Olpe, et al, Eur. J. Pharmacol., 187, 27 (1990); phosphinic acidderivatives described in Hills, et al, Br. J. Pharmacol., 102, pp. 5-6(1991); and compounds described in U.S. Pat. Nos. 4,656,298, 5,929,236,EP0463969, EP 0356128, Kaupmann et al., Nature 368: 239 (1997), Karla etal., J Med Chem., 42(11):2053-9 (1992), Ansar et al., Therapie,54(5):651-8 (1999), and Castelli et al., Eur J Pharmacol., 446(1-3): 1-5(2002).

In some embodiments, the GABA modulator modulates GABA-C receptoractivity. Non-limiting examples of reported GABA-C receptor modulatorsuseful in methods described herein include cis-aminocrotonic acid(CACA); 1,2,5,6-tetrahydropyridine-4-yl methyl phosphinic acid (TPMPA)and related compounds such as P4MPA, PPA and SEPI; 2-methyl-TACA;(+/−)-TAMP; muscimol and compounds disclosed in U.S. Pat. No. 3,242,190;ZAPA; THIP and related analogues, such as aza-THIP; pricotroxin;imidazole-4-acetic acid (IMA); and CGP36742.

In some embodiments, the GABA modulator modulates the activity ofglutamic acid decarboxylase (GAD).

In some embodiments, the GABA modulator modulates GABA transaminase(GTA). Non-limiting examples of GTA modulators include the GABA analogvigabatrin, and compounds disclosed in U.S. Pat. No. 3,960,927.

In some embodiments, the GABA modulator modulates the reuptake and/ortransport of GABA from extracellular regions. In other embodiments, theGABA modulator modulates the activity of the GABA transporters, GAT-1,GAT-2, GAT-3 and/or BGT-1. Non-limiting examples of GABA reuptake and/ortransport modulators include nipecotic acid and related derivatives,such as CI-966; SKF 89976A; TACA; stiripentol; tiagabine and GAT-1inhibitors disclosed in U.S. Pat. No. 5,010,090;(R)-1-(4,4-diphenyl-3-butenyl)-3-piperidinecarboxylic acid and relatedcompounds disclosed in U.S. Pat. No. 4,383,999;(R)-1-[4,4-bis(3-methyl-2-thienyl)-3-butenyl]-3-piperidinecarboxylicacid and related compounds disclosed in Anderson et al., J. Med. Chem.36, (1993) 1716-1725; guvacine and related compounds disclosed inKrogsgaard-Larsen, Molecular & Cellular Biochemistry 31, 105-121 (1980);GAT-4 inhibitors disclosed in U.S. Pat. No. 6,071,932; and compoundsdisclosed in 6,906,177 and Ali, F. E., et al. J. Med. Chem. 1985, 28,653-660. Methods for detecting GABA reuptake inhibitors are known in theart, and are described, e.g., in U.S. Pat. Nos. 6,906,177; 6,225,115;4,383,999; Ali, F. E., et al. J. Med. Chem. 1985, 28, 653-660.

In some embodiments, the GABA modulator is the benzodiazepineclonazepam, which is described, e.g., in U.S. Pat. Nos. 3,121,076 and3,116,203; the benzodiazepine diazepam, which is described, e.g., inU.S. Pat. Nos. 3,371,085; 3,109,843; and 3,136,815; the short-actingdiazepam derivative midazolam, which is a described, e.g., in U.S. Pat.No. 4,280,957; the imidazodiazepine flumazenil, which is described,e.g., in U.S. Pat. No. 4,316,839; the benzodiazepine lorazepam isdescribed, e.g., in U.S. Pat. No. 3,296,249; the benzodiazepineL-655708, which is described, e.g., in Quirk et al. Neuropharmacology1996, 35, 1331; Sur et al. Mol. Pharmacol. 1998, 54, 928; and Sur et al.Brain Res. 1999, 822, 265; the benzodiazepine gabitril; zopiclone, whichbinds the benzodiazepine site on GABA-A receptors, and is disclosed,e.g., in U.S. Pat. Nos. 3,862,149 and 4,220,646; the GABA-A potentiatorindiplon as described, e.g., in Foster et al., J Pharmacol Exp Ther.,311(2):547-59 (2004), U.S. Pat. Nos. 4,521,422 and 4,900,836; zolpidem,described, e.g., in U.S. Pat. No. 4,794,185 and EP50563; zaleplon,described, e.g., in U.S. Pat. No. 4,626,538; abecarnil, described, e.g.,in Stephens et al., J Pharmacol Exp Ther., 253(1):334-43 (1990); theGABA-A agonist isoguvacine, which is described, e.g., in Chebib et al.,Clin. Exp. Pharmacol. Physiol. 1999, 26, 937-940; Leinekugel et al. J.Physiol. 1995, 487, 319-29; and White et al., J. Neurochem. 1983, 40(6),1701-8; the GABA-A agonist gaboxadol (THIP), which is described, e.g.,in U.S. Pat. No. 4,278,676 and Krogsgaard-Larsen, Acta. Chem. Scand.1977, 31, 584; the GABA-A agonist muscimol, which is described, e.g., inU.S. Pat. Nos. 3,242,190 and 3,397,209; the inverse GABA-A agonistbeta-CCP, which is described, e.g., in Nielsen et al., J. Neurochem.,36(1):276-85 (1981); the GABA-A potentiator riluzole, which isdescribed, e.g., in U.S. Pat. No. 4,370,338 and EP 50,551; the GABA-Bagonist and GABA-C antagonist SKF 97541, which is described, e.g., inFroestl et al., J. Med. Chem. 38 3297 (1995); Hoskison et al., Neurosci.Lett. 2004, 365(1), 48-53 and Hue et al., J. Insect Physiol. 1997,43(12), 1125-1131; the GABA-B agonist baclofen, which is described,e.g., in U.S. Pat. No. 3,471,548; the GABA-C agonist cis-4-aminocrotonicacid (CACA), which is described, e.g., in Ulloor et al. J. Neurophysiol.2004, 91(4), 1822-31; the GABA-A antagonist phaclofen, which isdescribed, e.g., in Kerr et al. Brain Res. 1987, 405, 150; Karlsson etal. Eur. J. Pharmacol. 1988, 148, 485; and Hasuo, Gallagher Neurosci.Lett. 1988, 86, 77; the GABA-A antagonist SR 95531, which is described,e.g., in Stell et al. J. Neurosci. 2002, 22(10), RC223; Wermuth et al.,J. Med. Chem. 30 239 (1987); and Luddens and Korpi, J. Neurosci. 15:6957 (1995); the GABA-A antagonist bicuculline, which is a described,e.g., in Groenewoud, J. Chem. Soc. 1936, 199; Olsen et al., Brain Res.102: 283 (1976) and Haworth et al. Nature 1950, 165, 529; the selectiveGABA-B antagonist CGP 35348, which is described, e.g., in Olpe et al.Eur. J. Pharmacol. 1990, 187, 27; Hao et al. Neurosci. Lett. 1994, 182,299; and Froestl et al. Pharmacol. Rev. Comm. 1996, 8, 127; theselective GABA-B antagonist CGP 46381, which is described, e.g., inLingenhoehl, Pharmacol. Comm. 1993, 3, 49; the selective GABA-Bantagonist CGP 52432, which is described, e.g., in Lanza et al. Eur. J.Pharmacol. 1993, 237, 191; Froestl et al. Pharmacol. Rev. Comm. 1996, 8,127; Bonanno et al. Eur. J. Pharmacol. 1998, 362, 143; and Libri et al.Naunyn-Schmied. Arch. Pharmacol. 1998, 358, 168; the selective GABA-Bantagonist CGP 54626, which is described, e.g., in Brugger et al. Eur.J. Pharmacol. 1993, 235, 153; Froestl et al. Pharmacol. Rev. Comm. 1996,8, 127; and Kaupmann et al. Nature 1998, 396, 683; the selective GABA-Bantagonist CGP 55845, which is a GABA-receptor antagonist described,e.g., in Davies et al. Neuropharmacology 1993, 32, 1071; Froestl et al.Pharmacol. Rev. Comm. 1996, 8, 127; and Deisz Neuroscience 1999, 93,1241; the selective GABA-B antagonist Saclofen, which is described,e.g., in Bowery, TiPS, 1989, 10, 401; and Kerr et al. Neurosci Lett.1988; 92(1):92-6; the GABA-B antagonist 2-hydroxysaclofen, which isdescribed, e.g., in Kerr et al. Neurosci. Lett. 1988, 92, 92; and Curtiset al. Neurosci. Lett. 1988, 92, 97; the GABA-B antagonist SCH 50,911,which is described, e.g., in Carruthers et al., Bioorg Med Chem Lett 8:3059-3064 (1998); Bolser et al. J. Pharmacol. Exp. Ther. 1996, 274,1393; Hosford et al. J. Pharmacol. Exp. Ther. 1996, 274, 1399; and Onget al. Eur. J. Pharmacol. 1998, 362, 35; the selective GABA-C antagonistTPMPA, which is described, e.g., in Schlicker et al., Brain Res. Bull.2004, 63(2), 91-7; Murata et al., Bioorg. Med. Chem. Lett. 6: 2073(1996); and Ragozzino et al., Mol. Pharmacol. 50: 1024 (1996); a GABAderivative, such as Pregabalin [(S)-(+)-3-isobutylgaba] or gabapentin[1-(aminomethyl)cyclohexane acetic acid]. Gabapentin is described, e.g.,in U.S. Pat. No. 4,024,175; the lipid-soluble GABA agonist progabide,which is metabolized in vivo into GABA and/or pharmaceutically activeGABA derivatives in vivo. Progabide is described, e.g., in U.S. Pat.Nos. 4,094,992 and 4,361,583; the GAT1 inhibitor Tiagabine, which isdescribed, e.g., in U.S. Pat. No. 5,010,090 and Andersen et al. J. Med.Chem. 1993, 36, 1716; the GABA transaminase inhibitor valproic acid(2-propylpentanoic acid or dispropylacetic acid), which is described,e.g., in U.S. Pat. No. 4,699,927 and Carraz et al., Therapie, 1965, 20,419; the GABA transaminase inhibitor vigabatrin, which is described,e.g., in U.S. Pat. No. 3,960,927; or topiramate, which is described,e.g., in U.S. Pat. No. 4,513,006.

Additionally, the neurogenic agent in combination with an angiotensinagent may be a neurogenic sensitizing agent that is a reportedanti-epileptic agent. Non-limiting examples of such agents includecarbamazepine or tegretol (CAS RN 298-46-4), clonazepam (CAS RN1622-61-3), BPA or 3-(p-boronophenyl)alanine (CAS RN 90580-64-6),gabapentin or neurontin (CAS RN 60142-96-3), phenyloin (CAS RN 57-41-0),topiramate, lamotrigine or lamictal (CAS RN 84057-84-1), phenobarbital(CAS RN 50-06-6), oxcarbazepine (CAS RN 28721-07-5), primidone (CAS RN125-33-7), ethosuximide (CAS RN 77-67-8), levetiracetam (CAS RN102767-28-2), zonisamide, tiagabine (CAS RN 115103-54-3), depakote ordivalproex sodium (CAS RN 76584-70-8), felbamate (Na-channel and NMDAreceptor antagonist), or pregabalin (CAS RN 148553-50-8).

In further embodiments, the neurogenic sensitizing agent may be areported direct or indirect modulator of dopamine receptors.Non-limiting examples of such agents include the indirect dopamineagonists methylphenidate (CAS RN 113-45-1) or methylphenidatehydrochloride (also known as Ritalin® CAS RN 298-59-9), amphetamine (CASRN 300-62-9) and methamphetamine (CAS RN 537-46-2), and the directdopamine agonists sumanirole (CAS RN 179386-43-7), roprinirole (CAS RN91374-21-9), and rotigotine (CAS RN 99755-59-6). Additional non-limitingexamples include 7-OH-DPAT, quinpirole, haloperidole, or clozapine.

Additional non-limiting examples include bromocriptine (CAS RN25614-03-3), adrogolide (CAS RN 171752-56-0), pramipexole (CAS RN104632-26-0), ropinirole (CAS RN 91374-21-9), apomorphine (CAS RN58-00-4) or apomorphine hydrochloride (CAS RN 314-19-2), lisuride (CASRN 18016-80-3), sibenadet hydrochloride or viozan (CAS RN 154189-24-9),L-DOPA or levodopa (CAS RN 59-92-7), melevodopa (CAS RN 7101-51-1),etilevodopa (CAS RN 37178-37-3), talipexole hydrochloride (CAS RN36085-73-1) or talipexole (CAS RN 101626-70-4), nolomirole (CAS RN90060-42-7), quinelorane (CAS RN 97466-90-5), pergolide (CAS RN66104-22-1), fenoldopam (CAS RN 67227-56-9), carmoxirole (CAS RN98323-83-2), terguride (CAS RN 37686-84-3), cabergoline (CAS RN81409-90-7), quinagolide (CAS RN 87056-78-8) or quinagolidehydrochloride (CAS RN 94424-50-7), sumanirole, docarpamine (CAS RN74639-40-0), SLV-308 or 2(3H)-benzoxazolone,7-(4-methyl-1-piperazinyl)-monohydrochloride (CAS RN 269718-83-4),aripiprazole (CAS RN 129722-12-9), bifeprunox, lisdexamfetaminedimesylate (CAS RN 608137-33-3), safinamide (CAS RN 133865-89-1), oradderall or amfetamine (CAS RN 300-62-9).

In further embodiments, the neurogenic agent used in combination with anangiotensin agent may be a reported dual sodium and calcium channelmodulator. Non-limiting examples of such agents include safinamide andzonisamide. Additional non-limiting examples include enecadin (CAS RN259525-01-4), levosemotiadil (CAS RN 116476-16-5), bisaramil (CAS RN89194-77-4), SL-34.0829 (see U.S. Pat. No. 6,897,305), lifarizine (CASRN 119514-66-8), JTV-519(4-[3-(4-benzylpiperidin-1-yl)propionyl]-7-methoxy-2,3,4,5-tetrahydro-1,4-benzothiazepinemonohydrochloride), and delapril.

In further embodiments, the neurogenic agent in used in combination withan angiotensin agent may be a reported calcium channel antagonist suchas amlodipine (CAS RN 88150-42-9) or amlodipine maleate (CAS RN88150-47-4), nifedipine (CAS RN 21829-25-4), MEM-1003 (CAS RN see Roseet al. “Efficacy of MEM 1003, a novel calcium channel blocker, in delayand trace eyeblink conditioning in older rabbits.” Neurobiol Aging. 2006Apr. 16; [Epub ahead of print]), isradipine (CAS RN 75695-93-1),felodipine (CAS RN 72509-76-3; 3,5-Pyridinedicarboxylic acid,1,4-dihydro-4-(2,3-dichlorophenyl)-2,6-dimethyl-, ethyl methyl ester) orfelodipine (CAS RN 86189-69-7; 3,5-Pyridinedicarboxylic acid,4-(2,3-dichlorophenyl)-1,4-dihydro-2,6-dimethyl-, ethyl methyl ester,(+−)-), lemildipine (CAS RN 125729-29-5 or 94739-29-4), clevidipine (CASRN 166432-28-6 or 167221-71-8), verapamil (CAS RN 52-53-9), ziconotide(CAS RN 107452-89-1), monatepil maleate (CAS RN 132046-06-1), manidipine(CAS RN 89226-50-6), furnidipine (CAS RN 138661-03-7), nitrendipine (CASRN 39562-70-4), loperamide (CAS RN 53179-11-6), amiodarone (CAS RN1951-25-3), bepridil (CAS RN 64706-54-3), diltiazem (CAS RN 42399-41-7),nimodipine (CAS RN 66085-59-4), lamotrigine, cinnarizine (CAS RN298-57-7), lacipidine (CAS RN 103890-78-4), nilvadipine (CAS RN75530-68-6), dotarizine (CAS RN 84625-59-2), cilnidipine (CAS RN132203-70-4), oxodipine (CAS RN 90729-41-2), aranidipine (CAS RN86780-90-7), anipamil (CAS RN 83200-10-6), ipenoxazone (CAS RN104454-71-9), efonidipine hydrochloride or NZ 105 (CAS RN 111011-53-1)or efonidipine (CAS RN 111011-63-3), temiverine (CAS RN 173324-94-2),pranidipine (CAS RN 99522-79-9), dopropidil (CAS RN 79700-61-1),lercanidipine (CAS RN 100427-26-7), terodiline (CAS RN 15793-40-5),fantofarone (CAS RN 114432-13-2), azelnidipine (CAS RN 123524-52-7),mibefradil (CAS RN 116644-53-2) or mibefradil dihydrochloride (CAS RN116666-63-8), SB-237376 (see Xu et al. “Electrophysiologic effects ofSB-237376: a new antiarrhythmic compound with dual potassium and calciumchannel blocking action.” J Cardiovasc Pharmacol. 2003 41(3):414-21),BRL-32872 (CAS RN 113241-47-7), S-2150 (see Ishibashi et al.“Pharmacodynamics of S-2150, a simultaneous calcium-blocking andalpha1-inhibiting antihypertensive drug, in rats.” J Pharm Pharmacol.2000 52(3):273-80), nisoldipine (CAS RN 63675-72-9), semotiadil (CAS RN116476-13-2), palonidipine (CAS RN 96515-73-0) or palonidipinehydrochloride (CAS RN 96515-74-1), SL-87.0495 (see U.S. Pat. No.6,897,305), YM430 (4(((S)-2-hydroxy-3-phenoxypropyl)amino)butyl methyl2,6-dimethyl-((S)-4-(m-nitrophenyl))-1,4-dihydropyridine-3,5-dicarboxylate),barnidipine (CAS RN 104713-75-9), and AM336 or CVID (see Adams et al.“Omega-Conotoxin CVID Inhibits a Pharmacologically DistinctVoltage-sensitive Calcium Channel Associated with Transmitter Releasefrom Preganglionic Nerve Terminals” J. Biol. Chem., 278(6):4057-4062,2003). An additional non-limiting example is NMED-160.

In other embodiments, the neurogenic agent used in combination with anangiotensin agent may be a reported modulator of a melatonin receptor.Non-limiting examples of such modulators include the melatonin receptoragonists melatonin, LY-156735 (CAS RN 118702-11-7), agomelatine (CAS RN138112-76-2), 6-chloromelatonin (CAS RN 63762-74-3), ramelteon (CAS RN196597-26-9), 2-Methyl-6,7-dichloromelatonin (CAS RN 104513-29-3), andML 23 (CAS RN 108929-03-9).

In yet further embodiments, the neurogenic agent in combination with anangiotensin agent may be a reported modulator of a melanocortinreceptor. Non-limiting examples of such agents include a melanocortinreceptor agonists selected from melanotan II (CAS RN 121062-08-6),PT-141 or bremelanotide (CAS RN 189691-06-3), HP-228 (see Getting et al.“The melanocortin peptide HP228 displays protective effects in acutemodels of inflammation and organ damage.” Eur J Pharmacol. 2006 Jan.24), or AP214 from Action Pharma A/S.

Additionally, the agent used in combination with an angiotensin agentmay be a reported compound (or “monoamine modulator”) that modulatesneurotransmission mediated by one or more monoamine neurotransmitters(referred to herein as “monoamines”) or other biogenic amines, such astrace amines (TAs) as a non-limiting example. TAs are endogenous,CNS-active amines that are structurally related to classical biogenicamines (e.g., norepinephrine, dopamine(4-(2-aminoethyl)benzene-1,2-diol), and/or serotonin(5-hydroxytryptamine (5-HT), or a metabolite, precursor, prodrug, oranalog thereof. The methods of the disclosure thus includeadministration of one or more reported TAs in a combination with anangiotensin agent. Additional CNS-active monoamine receptor modulatorsare well known in the art, and are described, e.g., in the Merck Index,12th Ed. (1996).

Certain food products, e.g., chocolates, cheeses, and wines, can alsoprovide a significant dietary source of TAs and/or TA-related compounds.Non-limiting examples of mammalian TAs useful as constitutive factorsinclude, but are not limited to, tryptamine, ρ-tyramine, m-tyramine,octopamine, synephrine or β-phenylethylamine (β-PEA). Additional usefulTA-related compounds include, but are not limited to,5-hydroxytryptamine, amphetamine, bufotenin, 5-methoxytryptamine,dihydromethoxytryptamine, phenylephrine, or a metabolite, precursor,prodrug, or analogue thereof.

In some embodiments, the constitutive factor is a biogenic amine or aligand of a trace amine-associated receptor (TAAR), and/or an agent thatmediates one or more biological effects of a TA. TAs have been shown tobind to and activate a number of unique receptors, termed TAARs, whichcomprise a family of G-protein coupled receptors (TAAR1-TAAR9) withhomology to classical biogenic amine receptors. For example, TAAR1 isactivated by both tyramine and β-PEA.

Thus non-limiting embodiments include methods and combinationcompositions wherein the constitutive factor is β-PEA, which has beenindicated as having a significant neuromodulatory role in the mammalianCNS and is found at relatively high levels in the hippocampus (e.g.,Taga et al., Biomed Chromatogr., 3(3): 118-20 (1989)); a metabolite,prodrug, precursor, or other analogue of β-PEA, such as the β-PEAprecursor L-phenylalanine, the β-PEA metabolite β-phenylacetic acid(β-PAA), or the β-PEA analogues methylphenidate, amphetamine, andrelated compounds.

Most TAs and monoamines have a short half-life (e.g., less than about 30s) due, e.g., to their rapid extracellular metabolism. Thus embodimentsof the disclosure include use of a monoamine “metabolic modulator,”which increases the extracellular concentration of one or moremonoamines by inhibiting monoamine metabolism. In some embodiments, themetabolic modulator is an inhibitor of the enzyme monoamine oxidase(MAO), which catalyzes the extracellular breakdown of monoamines intoinactive species. Isoforms MAO-A and/or MAO-B provide the major pathwayfor TA metabolism. Thus, in some embodiments, TA levels are regulated bymodulating the activity of MAO-A and/or MAO-B. For example, in someembodiments, endogenous TA levels are increased (and TA signaling isenhanced) by administering an inhibitor of MAO-A and/or MAO-B, incombination with an angiotensin agent as described herein.

Non-limiting examples of inhibitors of monoamine oxidase (MAO) includereported inhibitors of the MAO-A isoform, which preferentiallydeaminates 5-hydroxytryptamine (serotonin) (5-HT) and norepinephrine(NE), and/or the MAO-B isoform, which preferentially deaminatesphenylethylamine (PEA) and benzylamine (both MAO-A and MAO-B metabolizeDopamine (DA)). In various embodiments, MAO inhibitors may beirreversible or reversible (e.g., reversible inhibitors of MAO-A(RIMA)), and may have varying potencies against MAO-A and/or MAO-B(e.g., non-selective dual inhibitors or isoform-selective inhibitors).Non-limiting examples of MAO inhibitors useful in methods describedherein include clorgyline, L-deprenyl, isocarboxazid (Marplan®),ayahuasca, nialamide, iproniazide, iproclozide, moclobemide (Aurorix®),phenelzine (Nardil®), tranylcypromine (Parnate®) (the congeneric ofphenelzine), toloxatone, levo-deprenyl (Selegiline®), harmala, RIMAs(e.g., moclobemide, described in Da Prada et al., J Pharmacol Exp Ther248: 400-414 (1989); brofaromine; and befloxatone, described in Curet etal., J Affect Disord 51: 287-303 (1998)), lazabemide (Ro 19 6327),described in Ann. Neurol., 40(1): 99-107 (1996), and SL25.1131,described in Aubin et al., J. Pharmacol. Exp. Ther., 310: 1171-1182(2004).

In additional embodiments, the monoamine modulator is an “uptakeinhibitor,” which increases extracellular monoamine levels by inhibitingthe transport of monoamines away from the synaptic cleft and/or otherextracellular regions. In some embodiments, the monoamine modulator is amonoamine uptake inhibitor, which may selectively/preferentially inhibituptake of one or more monoamines relative to one or more othermonoamines. The term “uptake inhibitors” includes compounds that inhibitthe transport of monoamines (e.g., uptake inhibitors) and/or the bindingof monoamine substrates (e.g., uptake blockers) by transporter proteins(e.g., the dopamine transporter (DAT), the NE transporter (NET), the5-HT transporter (SERT), and/or the extraneuronal monoamine transporter(EMT)) and/or other molecules that mediate the removal of extracellularmonoamines. Monoamine uptake inhibitors are generally classifiedaccording to their potencies with respect to particular monoamines, asdescribed, e.g., in Koe, J. Pharmacol. Exp. Ther. 199: 649-661 (1976).However, references to compounds as being active against one or moremonoamines are not intended to be exhaustive or inclusive of themonoamines modulated in vivo, but rather as general guidance for theskilled practitioner in selecting compounds for use in therapeuticmethods provided herein.

In embodiments relating to a biogenic amine modulator used in acombination or method with an angiotensin agent as disclosed herein, themodulator may be (i) a norepinephrine and dopamine reuptake inhibitor,such as bupropion (described, e.g., in U.S. Pat. Nos. 3,819,706 and3,885,046), or (S,S)-hydroxybupropion (described, e.g., in U.S. Pat. No.6,342,496); (ii) selective dopamine reuptake inhibitors, such asmedifoxamine, amineptine (described, e.g., in U.S. Pat. Nos. 3,758,528and 3,821,249), GBR12909, GBR12783 and GBR13069, described in Andersen,Eur J Pharmacol, 166:493-504 (1989); or (iii) a monoamine “releaser”which stimulates the release of monoamines, such as biogenic amines frompresynaptic sites, e.g., by modulating presynaptic receptors (e.g.,autoreceptors, heteroreceptors), modulating the packaging (e.g.,vesicular formation) and/or release (e.g., vesicular fusion and release)of monoamines, and/or otherwise modulating monoamine release.Advantageously, monoamine releasers provide a method for increasinglevels of one or more monoamines within the synaptic cleft or otherextracellular region independently of the activity of the presynapticneuron.

Monoamine releasers useful in combinations provided herein includefenfluramine or p-chloro amphetamine (PCA) or the dopamine,norepinephrine, and serotonin releasing compound amineptine (described,e.g., in U.S. Pat. Nos. 3,758,528 and 3,821,249).

The agent used with an angiotensin agent may be a reportedphosphodiesterase (PDE) inhibitor. In some embodiments, a reportedinhibitor of PDE activity include an inhibitor of a cAMP-specific PDE.Non-limiting examples of cAMP specific PDE inhibitors useful in themethods described herein include a pyrrolidinone, such as a compounddisclosed in U.S. Pat. No. 5,665,754, US20040152754 or US20040023945; aquinazolineone, such as a compound disclosed in U.S. Pat. Nos. 6,747,035or 6,828,315, WO 97/49702 or WO 97/42174; a xanthine derivative; aphenylpyridine, such as a compound disclosed in U.S. Pat. Nos. 6,410,547or 6,090,817, or WO 97/22585; a diazepine derivative, such as a compounddisclosed in WO 97/36905; an oxime derivative, such as a compounddisclosed in U.S. Pat. No. 5,693,659 or WO 96/00215; a naphthyridine,such as a compound described in U.S. Pat. Nos. 5,817,670, 6,740,662,6,136,821, 6,331,548, 6,297,248, 6,541,480, 6,642,250, or 6,900,205, orTrifilieff et al., Pharmacology, 301(1): 241-248 (2002), or Herspergeret al., J Med Chem., 43(4):675-82 (2000); a benzofuran, such as acompound disclosed in U.S. Pat. Nos. 5,902,824, 6,211,203, 6,514,996,6,716,987, 6,376,535, 6,080,782, or 6,054,475, or EP 819688, EP685479,or Perrier et al., Bioorg. Med. Chem. Lett. 9:323-326 (1999); aphenanthridine, such as that disclosed in U.S. Pat. Nos. 6,191,138,6,121,279, or 6,127,378; a benzoxazole, such as that disclosed in U.S.Pat. Nos. 6,166,041 or 6,376,485; a purine derivative, such as acompound disclosed in U.S. Pat. No. 6,228,859; a benzamide, such as acompound described in U.S. Pat. Nos. 5,981,527 or 5,712,298, orWO95/01338, WO 97/48697 or Ashton et al., J. Med Chem 37: 1696-1703(1994); a substituted phenyl compound, such as a compound disclosed inU.S. Pat. Nos. 6,297,264, 5,866,593,65 5,859,034, 6,245,774, 6,197,792,6,080,790, 6,077,854, 5,962,483, 5,674,880, 5,786,354, 5,739,144,5,776,958, 5,798,373, 5,891,896, 5,849,770, 5,550,137, 5,340,827,5,780,478, 5,780,477, or 5,633,257, or WO 95/35283; a substitutedbiphenyl compound, such as that disclosed in U.S. Pat. No. 5,877,190; ora quinilinone, such as a compound described in U.S. Pat. No. 6,800,625or WO 98/14432.

Additional non-limiting examples of reported cAMP-specific PDEinhibitors useful in methods disclosed herein include a compounddisclosed in U.S. Pat. Nos. 6,818,651, 6,737,436, 6,613,778, 6,617,357,6,146,876, 6,838,559, 6,884,800, 6,716,987, 6,514,996, 6,376,535,6,740,655, 6,559,168, 6,069,151, 6,365,585, 6,313,116, 6,245,774,6,011,037, 6,127,363, 6,303,789, 6,316,472, 6,348,602, 6,331,543,6,333,354, 5,491,147, 5,608,070, 5,622,977, 5,580,888, 6,680,336,6,569,890, 6,569,885, 6,500,856, 6,486,186, 6,458,787, 6,455,562,6,444,671, 6,423,710, 6,376,489, 6,372,777, 6,362,213, 6,313,156,6,294,561, 6,258,843, 6,258,833, 6,121,279, 6,043,263, RE38,624,6,297,257, 6,251,923, 6,613,794, 6,407,108, 6,107,295, 6,103,718,6,479,494, 6,602,890, 6,545,158, 6,545,025, 6,498,160, 6,743,802,6,787,554, 6,828,333, 6,869,945, 6,894,041, 6,924,292, 6,949,573,6,953,810, 6,156,753, 5,972,927, 5,962,492, 5,814,651, 5,723,460,5,716,967, 5,686,434, 5,502,072, 5,116,837, 5,091,431; 4,670,434;4,490,371; 5,710,160, 5,710,170, 6,384,236, or 3,941,785, orUS20050119225, US20050026913, US20050059686, US20040138279,US20050222138, US20040214843, US20040106631, US 20030045557, US20020198198, US20030162802, US20030092908, US 20030104974,US20030100571, 20030092721, US20050148604, WO 99/65880, WO 00/26201, WO98/06704, WO 00/59890, WO9907704, WO9422852, WO 98/20007, WO 02/096423,WO 98/18796, WO 98/02440, WO 02/096463, WO 97/44337, WO 97/44036, WO97/44322, EP 0763534, Aoki et al., J Pharmacol Exp Ther., 295(1):255-60(2000), Del Piaz et al., Eur. J. Med. Chem., 35; 463-480 (2000), orBarnette et al., Pharmacol. Rev. Commun. 8: 65-73 (1997).

In some embodiments, the reported cAMP-specific PDE inhibitor iscilomilast (SB-207499); filaminast; tibenelast (LY-186655); ibudilast;piclamilast (RP 73401); theophylline, doxofylline; cipamfylline(HEP-688); atizoram (CP-80633); isobutylmethylxanthine; mesopram(ZK-117137); zardaverine; vinpocetine; rolipram (ZK-62711); arofylline(LAS-31025); roflumilast (BY-217); pumafentrin (BY-343); denbufylline;EHNA; milrinone; siguazodan; zaprinast; tolafentrine; Isbufylline; IBMX;1C-485; dyphylline; verolylline; bamifylline; pentoxyfilline;enprofilline; lirimilast (BAY 19-8004); filaminast (WAY-PDA-641);benafentrine; trequinsin; nitroquazone; cilostamide; vesnarinone;piroximone; enoximone; aminone; olprinone; imazodan or5-methyl-imazodan; indolidan; anagrelide; carbazeran; ampizone;emoradan; motapizone; phthalazinol; lixazinone (RS 82856); quazinone;bemorandan (RWJ 22867); adibendan (BM 14,478); pimobendan (MCI-154);saterinone (BDF 8634); tetomilast (OPC-6535); benzafentrine; sulmazole(ARL 115); revizinone; 349-U-85; AH-21-132; ATZ-1993; AWD-12-343;AWD-12-281; AWD-12-232; BRL 50481; CC-7085; CDC-801; CDC-998; CDP-840;CH-422; CH-673; CH-928; CH-3697; CH-3442; CH-2874; CH-4139; Chiroscience245412; CI-930; CI-1018; CI-1044; CI-1118; CP-353164; CP-77059;CP-146523; CP-293321; CP-220629; CT-2450; CT-2820; CT-3883; CT-5210;D-4418; D-22888; E-4021; EMD 54622; EMD-53998; EMD-57033; GF-248;GW-3600; IC-485; ICI-63197; ICI 153,110; IPL-4088; KF-19514; KW-4490;L-787258; L-826141; L-791943; LY181512; NCS-613; NM-702; NSP-153;NSP-306; NSP-307; Org-30029; Org-20241; Org-9731; ORG 9935; PD-168787;PD-190749; PD-190036; PDB-093; PLX650; PLX369; PLX371; PLX788; PLX939;Ro-20-1724; RPR-132294; RPR-117658A; RPR-114597; RPR-122818; RPR-132703;RS-17597; RS-25344; RS-14203; SCA 40; Sch-351591; SDZ-ISQ-844;SDZ-MKS-492; SKF 94120; SKF-95654; SKF-107806; SKF 96231; T-440; T-2585;WAY-126120; WAY-122331; WAY-127093B; WIN-63291; WIN-62582; V-11294A; VMX554; VMX 565; XT-044; XT-611; Y-590; YM-58897; YM-976; ZK-62711; methyl3-[6-(2H-3,4,5,6-tetrahydropyran-2-yloxy)-2-(3-thienylcarbonyl)benzo[b]furan-3-yl]propanoate;4-[4-methoxy-3-(5-phenylpentyloxy)phenyl]-2-methylbenzoic acid; methyl3-{2-[(4-chlorophenyl)carbonyl]-6-hydroxybenzo[b]furan-3-yl}propanoate;(R*,R*)-(±)-methyl3-acetyl-4-[3-(cyclopentyloxy)-4-methoxyphenyl]-3-methyl-1-pyrrolidinecarboxylat;or 4-(3-bromophenyl)-1-ethyl-7-methylhydropyridino[2,3-b]pyridin-2-one.

In some embodiments, the reported PDE inhibitor inhibits a cGMP-specificPDE. Non-limiting examples of a cGMP specific PDE inhibitor for use inthe combinations and methods described herein include a pyrimidine orpyrimidinone derivative, such as a compound described in U.S. Pat. Nos.6,677,335, 6,458,951, 6,251,904, 6,787,548, 5,294,612, 5,250,534, or6,469,012, WO 94/28902, WO96/16657, EP0702555, and Eddahibi, Br. J.Pharmacol., 125(4): 681-688 (1988); a griseolic acid derivative, such asa compound disclosed in U.S. Pat. No. 4,460,765; a 1-arylnaphthalenelignan, such as that described in Ukita, J. Med. Chem. 42(7): 1293-1305(1999); a quinazoline derivative, such as4-[[3′,4′-(methylenedioxy)benzyl]amino]-6-methoxyquinazoline) or acompound described in U.S. Pat. Nos. 3,932,407 or 4,146,718, orRE31,617; a pyrroloquinolone or pyrrolopyridinone, such as thatdescribed in U.S. Pat. Nos. 6,686,349, 6,635,638, 6,818,646,US20050113402; a carboline derivative, such a compound described in U.S.Pat. Nos. 6,492,358, 6,462,047, 6,821,975, 6,306,870, 6,117,881,6,043,252, or 3,819,631, US20030166641, WO 97/43287, Daugan et al., JMed Chem., 46(21):4533-42 (2003), or Daugan et al., J Med Chem., 9;46(21):4525-32 (2003); an imidazo derivative, such as a compounddisclosed in U.S. Pat. Nos. 6,130,333, 6,566,360, 6,362,178, or6,582,351, US20050070541, or US20040067945; or a compound described inU.S. Pat. Nos. 6,825,197, 5,719,283, 6,943,166, 5,981,527, 6,576,644,5,859,009, 6,943,253, 6,864,253, 5,869,516, 5,488,055, 6,140,329,5,859,006, or 6,143,777, WO 96/16644, WO 01/19802, WO 96/26940, Dunn,Org. Proc. Res. Dev., 9: 88-97 (2005), or Bi et al., Bioorg Med ChemLett., 11(18):2461-4 (2001).

In some embodiments, the PDE inhibitor used in a combination or methoddisclosed herein is caffeine. In some embodiments, the caffeine isadministered in a formulation comprising an angiotensin agent. In otherembodiments, the caffeine is administered simultaneously with anangiotensin agent. In alternative embodiments, the caffeine isadministered in a formulation, dosage, or concentration lower or higherthan that of a caffeinated beverage such as coffee, tea, or soft drinks.In further embodiments, the caffeine is administered by a non-oralmeans, including, but not limited to, parenteral (e.g., intravenous,intradermal, subcutaneous, inhalation), transdermal (topical),transmucosal, rectal, or intranasal (including, but not limited to,inhalation of aerosol suspensions for delivery of compositions to thenasal mucosa, trachea and bronchioli) administration. The disclosureincludes embodiments with the explicit exclusion of caffeine or anotherone or more of the described agents for use in combination with anangiotensin agent.

In further alternative embodiments, the caffeine is in an isolated form,such as that which is separated from one or more molecules ormacromolecules normally found with caffeine before use in a combinationor method as disclosed herein. In other embodiments, the caffeine iscompletely or partially purified from one or more molecules ormacromolecules normally found with the caffeine. Exemplary cases ofmolecules or macromolecules found with caffeine include a plant or plantpart, an animal or animal part, and a food or beverage product.

Non-limiting examples of a reported PDE1 inhibitor include IBMX;vinpocetine; MMPX; KS-505a; SCH-51866; W-7; PLX650; PLX371; PLX788; aphenothiazines; or a compound described in U.S. Pat. No. 4,861,891.

Non-limiting examples of a PDE2 inhibitor include EHNA; PLX650; PLX369;PLX788; PLX 939; Bay 60-7550 or a related compound described in Boess etal., Neuropharmacology, 47(7): 1081-92 (2004); or a compound describedin US20020132754.

Non-limiting examples of reported PDE3 inhibitors include adihydroquinolinone compound such as cilostamide, cilostazol,vesnarinone, or OPC 3911; an imidazolone such as piroximone orenoximone; a bipyridine such as milrinone, aminone or olprinone; animidazoline such as imazodan or 5-methyl-imazodan; a pyridazinone suchas indolidan; LY181512 (see Komas et al. “Differential sensitivity tocardiotonic drugs of cyclic AMP phosphodiesterases isolated from canineventricular and sinoatrial-enriched tissues.” J Cardiovasc Pharmacol.1989 14(2):213-20); ibudilast; isomazole; motapizone; phthalazinol;trequinsin; lixazinone (RS 82856); Y-590; SKF 94120; quazinone; ICI153,110; bemorandan (RWJ 22867); siguazodan (SK&F 94836); adibendan (BM14,478); pimobendan (UD-CG 115, MCI-154); saterinone (BDF 8634);NSP-153; zardaverine; a quinazoline; benzafentrine; sulmazole (ARL 115);ORG 9935; CI-930; SKF-95654; SDZ-MKS-492; 349-U-85; EMD-53998;EMD-57033; NSP-306; NSP-307; Revizinone; NM-702; WIN-62582; ATZ-1993;WIN-63291; ZK-62711; PLX650; PLX369; PLX788; PLX939; anagrelide;carbazeran; ampizone; emoradan; or a compound disclosed in U.S. Pat. No.6,156,753.

Non-limiting examples of reported PDE4 inhibitors include apyrrolidinone, such as a compound disclosed in U.S. Pat. No. 5,665,754,US20040152754 or US20040023945; a quinazolineone, such as a compounddisclosed in U.S. Pat. No. 6,747,035 or 6,828,315, WO 97/49702 or WO97/42174; a xanthine derivative; a phenylpyridine, such as a compounddisclosed in U.S. Pat. No. 6,410,547 or 6,090,817 or WO 97/22585; adiazepine derivative, such as a compound disclosed in WO 97/36905; anoxime derivative, such as a compound disclosed in U.S. Pat. No.5,693,659 or WO 96/00215; a naphthyridine, such as a compound describedin U.S. Pat. Nos. 5,817,670, 6,740,662, 6,136,821, 6,331,548, 6,297,248,6,541,480, 6,642,250, or 6,900,205, Trifilieff et al., Pharmacology,301(1): 241-248 (2002) or Hersperger et al., J Med Chem., 43(4):675-82(2000); a benzofuran, such as a compound disclosed in U.S. Pat. Nos.5,902,824, 6,211,203, 6,514,996, 6,716,987, 6,376,535, 6,080,782, or6,054,475, EP 819688, EP685479, or Perrier et al., Bioorg. Med. Chem.Lett. 9:323-326 (1999); a phenanthridine, such as that disclosed in U.S.Pat. Nos. 6,191,138, 6,121,279, or 6,127,378; a benzoxazole, such asthat disclosed in U.S. Pat. Nos. 6,166,041 or 6,376,485; a purinederivative, such as a compound disclosed in U.S. Pat. No. 6,228,859; abenzamide, such as a compound described in U.S. Pat. No. 5,981,527 or5,712,298, WO95/01338, WO 97/48697, or Ashton et al., J. Med Chem 37:1696-1703 (1994); a substituted phenyl compound, such as a compounddisclosed in U.S. Pat. Nos. 6,297,264, 5,866,593,65 5,859,034,6,245,774, 6,197,792, 6,080,790, 6,077,854, 5,962,483, 5,674,880,5,786,354, 5,739,144, 5,776,958, 5,798,373, 5,891,896, 5,849,770,5,550,137, 5,340,827, 5,780,478, 5,780,477, or 5,633,257, or WO95/35283; a substituted biphenyl compound, such as that disclosed inU.S. Pat. No. 5,877,190; or a quinilinone, such as a compound describedin U.S. Pat. No. 6,800,625 or WO 98/14432.

Additional examples of reported PDE4 inhibitors useful in methodsprovided herein include a compound disclosed in U.S. Pat. Nos.6,716,987, 6,514,996, 6,376,535, 6,740,655, 6,559,168, 6,069,151,6,365,585, 6,313,116, 6,245,774, 6,011,037, 6,127,363, 6,303,789,6,316,472, 6,348,602, 6,331,543, 6,333,354, 5,491,147, 5,608,070,5,622,977, 5,580,888, 6,680,336, 6,569,890, 6,569,885, 6,500,856,6,486,186, 6,458,787, 6,455,562, 6,444,671, 6,423,710, 6,376,489,6,372,777, 6,362,213, 6,313,156, 6,294,561, 6,258,843, 6,258,833,6,121,279, 6,043,263, RE38,624, 6,297,257, 6,251,923, 6,613,794,6,407,108, 6,107,295, 6,103,718, 6,479,494, 6,602,890, 6,545,158,6,545,025, 6,498,160, 6,743,802, 6,787,554, 6,828,333, 6,869,945,6,894,041, 6,924,292, 6,949,573, 6,953,810, 5,972,927, 5,962,492,5,814,651, 5,723,460, 5,716,967, 5,686,434, 5,502,072, 5,116,837,5,091,431; 4,670,434; 4,490,371; 5,710,160, 5,710,170, 6,384,236, or3,941,785, US20050119225, US20050026913, WO 99/65880, WO 00/26201, WO98/06704, WO 00/59890, WO9907704, WO9422852, WO 98/20007, WO 02/096423,WO 98/18796, WO 98/02440, WO 02/096463, WO 97/44337, WO 97/44036, WO97/44322, EP 0763534, Aoki et al., J Pharmacol Exp Ther., 295(1):255-60(2000), Del Piaz et al., Eur. J. Med. Chem., 35; 463-480 (2000), orBarnette et al., Pharmacol. Rev. Commun. 8: 65-73 (1997).

Non-limiting examples of a reported PDE5 inhibitor useful in acombination or method described herein include a pyrimidine orpyrimidinone derivative, such as a compound described in U.S. Pat. Nos.6,677,335, 6,458,951, 6,251,904, 6,787,548, 5,294,612, 5,250,534, or6,469,012, WO 94/28902, WO96/16657, EP0702555, or Eddahibi, Br. J.Pharmacol., 125(4): 681-688 (1988); a griseolic acid derivative, such asa compound disclosed in U.S. Pat. No. 4,460,765; a 1-arylnaphthalenelignan, such as that described in Ukita, J. Med. Chem. 42(7): 1293-1305(1999); a quinazoline derivative, such as4-[[3′,4′-(methylenedioxy)benzyl]amino]-6-methoxyquinazoline) or acompound described in U.S. Pat. Nos. 3,932,407 or 4,146,718, orRE31,617; a pyrroloquinolones or pyrrolopyridinone, such as thatdescribed in U.S. Pat. Nos. 6,686,349, 6,635,638, or 6,818,646,US20050113402; a carboline derivative, such a compound described in U.S.Pat. Nos. 6,492,358, 6,462,047, 6,821,975, 6,306,870, 6,117,881,6,043,252, or 3,819,631, US20030166641, WO 97/43287, Daugan et al., JMed Chem., 46(21):4533-42 (2003), and Daugan et al., J Med Chem., 9;46(21):4525-32 (2003); an imidazo derivative, such as a compounddisclosed in U.S. Pat. Nos. 6,130,333, 6,566,360, 6,362,178, or6,582,351, US20050070541, or US20040067945; or a compound described inU.S. Pat. Nos. 6,825,197, 6,943,166, 5,981,527, 6,576,644, 5,859,009,6,943,253, 6,864,253, 5,869,516, 5,488,055, 6,140,329, 5,859,006, or6,143,777, WO 96/16644, WO 01/19802, WO 96/26940, Dunn, Org. Proc. Res.Dev., 9: 88-97 (2005), or Bi et al., Bioorg Med Chem Lett.,11(18):2461-4 (2001).

In some embodiments, a reported PDE5 inhibitor is zaprinast; MY-5445;dipyridamole; vinpocetine; FR229934;1-methyl-3-isobutyl-8-(methylamino)xanthine; furazlocillin; Sch-51866;E4021; GF-196960; IC-351; T-1032; sildenafil; tadalafil; vardenafil;DMPPO; RX-RA-69; KT-734; SKF-96231; ER-21355; BF/GP-385; NM-702; PLX650;PLX134; PLX369; PLX788; or vesnarinone.

In some embodiments, the reported PDE5 inhibitor is sildenafil or arelated compound disclosed in U.S. Pat. Nos. 5,346,901, 5,250,534, or6,469,012; tadalafil or a related compound disclosed in U.S. Pat. Nos.5,859,006, 6,140,329, 6,821,975, or 6,943,166; or vardenafil or arelated compound disclosed in U.S. Pat. No. 6,362,178.

Non-limiting examples of a reported PDE6 inhibitor useful in acombination or method described herein include dipyridamole orzaprinast.

Non-limiting examples of a reported PDE7 inhibitor for use in thecombinations and methods described herein include BRL 50481; PLX369;PLX788; or a compound described in U.S. Pat. Nos. 6,818,651; 6,737,436,6,613,778, 6,617,357; 6,146,876, 6,838,559, or 6,884,800, US20050059686;US20040138279; US20050222138; US20040214843; US20040106631; US20030045557; US 20020198198; US20030162802, US20030092908, US20030104974; US20030100571; 20030092721; or US20050148604.

A non-limiting examples of a reported inhibitor of PDE8 activity isdipyridamole.

Non-limiting examples of a reported PDE9 inhibitor useful in acombination or method described herein include SCH-51866; IBMX; or BAY73-6691.

Non-limiting examples of a PDE10 inhibitor include sildenafil;SCH-51866; papaverine; zaprinast; dipyridamole; E4021; vinpocetine;EHNA; milrinone; rolipram; PLX107; or a compound described in U.S. Pat.No. 6,930,114, US20040138249, or US20040249148.

Non-limiting examples of a PDE 11 inhibitor includes IC-351 or a relatedcompound described in WO 9519978; E4021 or a related compound describedin WO 9307124; UK-235,187 or a related compound described in EP 579496;PLX788; zaprinast; dipyridamole; or a compound described inUS20040106631 or Maw et al., Bioorg Med Chem Lett. 2003 Apr. 17;13(8):1425-8.

In some embodiments, the reported PDE inhibitor is a compound describedin U.S. Pat. Nos. 5,091,431, 5,081,242, 5,066,653, 5,010,086, 4,971,972,4,963,561, 4,943,573, 4,906,628, 4,861,891, 4,775,674, 4,766,118,4,761,416, 4,739,056, 4,721,784, 4,701,459, 4,670,434, 4,663,320,4,642,345, 4,593,029, 4,564,619, 4,490,371, 4,489,078, 4,404,380,4,370,328, 4,366,156, 4,298,734, 4,289,772, RE30,511, 4,188,391,4,123,534, 4,107,309, 4,107,307, 4,096,257, 4,093,617, 4,051,236, or4,036,840.

In some embodiments, the reported PDE inhibitor inhibitsdual-specificity PDE. Non-limiting examples of a dual-specificity PDEinhibitor useful in a combination or method described herein include acAMP-specific or cGMP-specific PDE inhibitor described herein; MMPX;KS-505a; W-7; a phenothiazine; Bay 60-7550 or a related compounddescribed in Boess et al., Neuropharmacology, 47(7):1081-92 (2004);UK-235,187 or a related compound described in EP 579496; or a compounddescribed in U.S. Pat. Nos. 6,930,114 or 4,861,891, US20020132754,US20040138249, US20040249148, US20040106631, WO 951997, or Maw et al.,Bioorg Med Chem Lett. 2003 Apr. 17; 13(8):1425-8.

In some embodiments, a reported PDE inhibitor exhibits dual-selectivity,being substantially more active against two PDE isozymes relative toother PDE isozymes. For example, in some embodiments, a reported PDEinhibitor is a dual PDE4/PDE7 inhibitor, such as a compound described inUS20030104974; a dual PDE3/PDE4 inhibitor, such as zardaverine,tolafentrine, benafentrine, trequinsine, Org-30029, L-686398,SDZ-ISQ-844, Org-20241, EMD-54622, or a compound described in U.S. Pat.Nos. 5,521,187, or 6,306,869; or a dual PDE1/PDE4 inhibitor, such asKF19514(5-phenyl-3-(3-pyridyl)methyl-3H-imidazo[4,5-c][1,8]naphthyridin-4(5H)-one).

Furthermore, the neurogenic agent in combination with an angiotensinagent may be a reported neurosteroid. Non-limiting examples of such aneurosteroid include pregnenolone and allopregnenalone.

Alternatively, the neurogenic sensitizing agent may be a reportednon-steroidal anti-inflammatory drug (NSAID) or an anti-inflammatorymechanism targeting agent in general. Non-limiting examples of areported NSAID include a cyclooxygenase inhibitor, such as indomethacin,ibuprofen, celecoxib, cofecoxib, naproxen, or aspirin. Additionalnon-limiting examples for use in combination with an angiotensin agentinclude rofecoxib, meloxicam, piroxicam, valdecoxib, parecoxib,etoricoxib, etodolac, nimesulide, acemetacin, bufexamac, diflunisal,ethenzamide, etofenamate, flobufen, isoxicam, kebuzone, lonazolac,meclofenamic acid, metamizol, mofebutazone, niflumic acid,oxyphenbutazone, paracetamol, phenidine, propacetamol, propyphenazone,salicylamide, tenoxicam, tiaprofenic acid, oxaprozin, lornoxicam,nabumetone, minocycline, benorylate, aloxiprin, salsalate, flurbiprofen,ketoprofen, fenoprofen, fenbufen, benoxaprofen, suprofen, piroxicam,meloxicam, diclofenac, ketorolac, fenclofenac, sulindac, tolmetin,xyphenbutazone, phenylbutazone, feprazone, azapropazone, flufenamic acidor mefenamic acid.

In additional embodiments, the neurogenic agent in combination with anangiotensin agent may be a reported agent for treating migraines.Non-limiting examples of such an agent include a triptan, such asalmotriptan or almotriptan malate; naratriptan or naratriptanhydrochloride; rizatriptan or rizatriptan benzoate; sumatriptan orsumatriptan succinate; zolmatriptan or zolmitriptan, frovatriptan orfrovatriptan succinate; or eletriptan or eletriptan hydrobromide.Embodiments of the disclosure may exclude combinations of triptans andan SSRI or SNRI that result in life threatening serotonin syndrome.

Other non-limiting examples include an ergot derivative, such asdihydroergotamine or dihydroergotamine mesylate, ergotamine orergotamine tartrate; diclofenac or diclofenac potassium or diclofenacsodium; flurbiprofen; amitriptyline; nortriptyline; divalproex ordivalproex sodium; propranolol or propranolol hydrochloride; verapamil;methysergide (CAS RN 361-37-5); metoclopramide; prochlorperazine (CAS RN58-38-8); acetaminophen; topiramate; GW274150([2-[(1-iminoethyl)amino]ethyl]-L-homocysteine); or ganaxalone (CAS RN38398-32-2).

Additional non-limiting examples include a COX-2 inhibitor, such ascelecoxib.

In other embodiments, the neurogenic agent in combination with anangiotensin agent may be a reported modulator of a nuclear hormonereceptor. Nuclear hormone receptors are activated via ligandinteractions to regulate gene expression, in some cases as part of cellsignaling pathways. Non-limiting examples of a reported modulatorinclude a dihydrotestosterone agonist such as dihydrotestosterone; a2-quinolone like LG121071(4-ethyl-1,2,3,4-tetrahydro-6-(trifluoromethyl)-8-pyridono[5,6-g]-quinoline);a non-steroidal agonist or partial agonist compound described in U.S.Pat. No. 6,017,924; LGD2226 (see WO 01/16108, WO 01/16133, WO 01/16139,and Rosen et al. “Novel, non-steroidal, selective androgen receptormodulators (SARMs) with anabolic activity in bone and muscle andimproved safety profile.” J Musculoskelet Neuronal Interact. 20022(3):222-4); or LGD2941 (from collaboration between LigandPharmaceuticals Inc. and TAP Pharmaceutical Products Inc.).

Additional non-limiting examples of a reported modulator include aselective androgen receptor modulator (SARM) such as andarine, ostarine,prostarin, or andromustine (all from GTx, Inc.); bicalutamide or abicalutamide derivative such as GTx-007 (U.S. Pat. No. 6,492,554); or aSARM as described in U.S. Pat. No. 6,492,554.

Further non-limiting examples of a reported modulator include anandrogen receptor antagonist such as cyproterone, bicalutamide,flutamide, or nilutamide; a 2-quinolone such as LG120907, represented bythe following structure:

or a derivative compound represented by the following structure:

(see Allan et al. “Therapeutic androgen receptor ligands” Nucl ReceptSignal 2003; 1: e009); a phthalamide, such as a modulator as describedby Miyachi et al. (“Potent novel nonsteroidal androgen antagonists witha phthalimide skeleton.” Bioorg. Med. Chem. Lett. 1997 7:1483-1488);osaterone or osaterone acetate; hydroxyflutamide; or a non-steroidalantagonist described in U.S. Pat. No. 6,017,924.

Other non-limiting examples of a reported modulator include a retinoicacid receptor agonist such as all-trans retinoic acid (Tretinoin®);isotretinoin (13-cis-retinoic acid); 9-cis retinoic acid; bexarotene;TAC-101 (4-[3,5-bis(trimethylsilyl)benzamide]benzoic acid); AC-261066(see Lund et al. “Discovery of a potent, orally available, andisoform-selective retinoic acid beta2 receptor agonist.” J Med Chem.2005 48(24):7517-9); LGD1550((2E,4E,6E)-3-methyl-7-(3,5-di-ter-butylphen-yl)octatrienoic acid);E6060 (E6060[4-{5-[7-fluoro-4-(trifluoromethyl)benzo[b]furan-2-yl]-1H-2-pyrrolyl}benzoicacid]; agonist 1 or 2 as described by Schapira et al. (“In silicodiscovery of novel Retinoic Acid Receptor agonist structures.” BMCStruct Biol. 2001; 1:1 (published online 2001 Jun. 4) where “Agonist 1was purchased from Bionet Research (catalog number 1G-433S). Agonist 2was purchased from Sigma-Aldrich (Sigma Aldrich library of rarechemicals. Catalog number S08503-1”); a synthetic acetylenic retinoicacid, such as AGN 190121 (CAS RN: 132032-67-8), AGN 190168 (ortazarotene or CAS RN 118292-40-3), or its metabolite AGN 190299 (CAS RN118292-41-4); etretinate; acitretin; an acetylenic retinoate, such asAGN 190073 (CAS 132032-68-9), or AGN 190089 (or 3-pyridinecarboxylicacid, 6-(4-(2,6,6-trimethyl-1-cyclohexen-1-yl)-3-buten-1-ynyl)-, ethylester or CAS RN 116627-73-7).

In further embodiments, the additional agent for use in combination withan angiotensin agent may be a reported modulator selected from thyroxin,tri-iodothyronine, or levothyroxine.

Alternatively, the additional agent is a vitamin D(1,25-dihydroxyvitamine D₃) receptor modulator, such as calcitriol or acompound described in Ma et al. (“Identification and characterization ofnoncalcemic, tissue-selective, nonsecosteroidal vitamin D receptormodulators.” J Clin Invest. 2006 116(4):892-904) or Molnar et al.(“Vitamin D receptor agonists specifically modulate the volume of theligand-binding pocket.” J Biol Chem. 2006 281(15):10516-26) or Millikenet al. (“EB1089, a vitamin D receptor agonist, reduces proliferation anddecreases tumor growth rate in a mouse model of hormone-induced mammarycancer.” Cancer Lett. 2005 229(2):205-15) or Yee et al. (“Vitamin Dreceptor modulators for inflammation and cancer.” Mini Rev Med. Chem.2005 5(8):761-78) or Adachi et al. “Selective activation of vitamin Dreceptor by lithocholic acid acetate, a bile acid derivative.” J LipidRes. 2005 46(1):46-57).

Furthermore, the additional agent may be a reported cortisol receptormodulator, such as methylprednisolone or its prodrug methylprednisolonesuleptanate; PI-1020 (NCX-1020 or budesonide-21-nitrooxymethylbenzoate);fluticasone furoate; GW-215864; betamethasone valerate; beclomethasone;prednisolone; or BVT-3498 (AMG-311).

Alternatively, the additional agent may be a reported aldosterone (ormineralocorticoid) receptor modulator, such as spironolactone oreplerenone.

In other embodiments, the additional agent may be a reportedprogesterone receptor modulator such as asoprisnil (CAS RN 199396-76-4);mesoprogestin or J1042; J956; medroxyprogesterone acetate (MPA); RS020;tanaproget; trimegestone; progesterone; norgestomet; melengestrolacetate; mifepristone; onapristone; ZK137316; ZK230211 (see Fuhrmann etal. “Synthesis and biological activity of a novel, highly potentprogesterone receptor antagonist.” J Med Chem. 2000 43(26):5010-6); or acompound described in Spitz “Progesterone antagonists and progesteronereceptor modulators: an overview.” Steroids 2003 68(10-13):981-93.

In further embodiments, the additional agent may be a reported i)peroxisome proliferator-activated receptor (PPAR) agonist such asmuraglitazar; tesaglitazar; reglitazar; GW-409544 (see Xu et al.“Structural determinants of ligand binding selectivity between theperoxisome proliferator-activated receptors.” Proc Natl Acad Sci USA.2001 98(24):13919-24); or DRL 11605 (Dr. Reddy's Laboratories); ii) aperoxisome proliferator-activated receptor alpha agonist likeclofibrate; ciprofibrate; fenofibrate; gemfibrozil; DRF-10945 (Dr.Reddy's Laboratories); iii) a peroxisome proliferator-activated receptordelta agonist such as GW501516 (CAS RN 317318-70-0); or iv) a peroxisomeproliferator-activated gamma receptor agonist like ahydroxyoctadecadienoic acid (HODE); (v) a prostaglandin derivative, suchas 15-deoxy-Delta12,14-prostaglandin J2; a thiazolidinedione(glitazone), such as pioglitazone, troglitazone; rosiglitazone orrosiglitazone maleate; ciglitazone; balaglitazone or DRF-2593; AMG 131(from Amgen); or G1262570 (from GlaxoWellcome). In additionalembodiments, a PPAR ligand is a PPARγ antagonist such as T0070907 (CASRN 313516-66-4) or GW9662 (CAS RN 22978-25-2).

In additional embodiments, the additional agent may be a reportedmodulator of an “orphan” nuclear hormone receptor. Embodiments include areported modulator of a liver X receptor, such as a compound describedin U.S. Pat. No. 6,924,311; a farnesoid X receptor, such as GW4064 asdescribed by Maloney et al. (“Identification of a chemical tool for theorphan nuclear receptor FXR.” J Med Chem. 2000 43(16):2971-4); a RXRreceptor; a CAR receptor, such as1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP); or a PXR receptor,such as SR-12813 (tetra-ethyl2-(3,5-di-tert-butyl-4-hydroxyphenyl)ethenyl-1,1-bisphosphonate).

In additional embodiments, the agent in combination with an angiotensinagent is ethyl eicosapentaenoate or ethyl-EPA (also known as5,8,11,14,17-eicosapentaenoic acid ethyl ester or miraxion, CAS RN86227-47-6), docosahexaenoic acid (DHA), or a retinoid acid drug. As anadditional non-limiting example, the agent may be omacor, a combinationof DHA and EPA, or idebenone (CAS RN 58186-27-9).

In further embodiments, a reported nootropic compound may be used as anagent in combination with an angiotensin agent. Non-limiting examples ofsuch a compound include piracetam (Nootropil®), aniracetam, xxiracetam,pramiracetam, pyritinol (Enerbol®), ergoloid mesylates (Hydergine®),galantamine or galantamine hydrobromide, selegiline, centrophenoxine(Lucidril®), desmopressin (DDAVP), nicergoline, vinpocetine, picamilon,vasopressin, milacemide, FK-960, FK-962, levetiracetam, nefiracetam, orhyperzine A (CAS RN: 102518-79-6).

Additional non-limiting examples of such a compound include anapsos (CASRN 75919-65-2), nebracetam (CAS RN 97205-34-0 or 116041-13-5),metrifonate, ensaculin (or CAS RN 155773-59-4 or KA-672) or ensaculinHCl, rokan (CAS RN 122933-57-7 or EGb 761), AC-3933(5-(3-methoxyphenyl)-3-(5-methyl-1,2,4-oxadiazol-3-yl)-2-oxo-1,2-dihydro-1,6-naphthyridine)or its hydroxylated metabolite SX-5745(3-(5-hydroxymethyl-1,2,4-oxadiazol-3-yl)-5-(3-methoxyphenyl)-2-oxo-1,2-dihydro-1,6-naphthyridine),JTP-2942 (CAS RN 148152-77-6), sabeluzole (CAS RN 104383-17-7),ladostigil (CAS RN 209394-27-4), choline alphoscerate (CAS RN 28319-77-9or Gliatilin®), dimebon (CAS RN 3613-73-8), tramiprosate (CAS RN3687-18-1), omigapil (CAS RN 181296-84-4), cebaracetam (CAS RN113957-09-8), fasoracetam (CAS RN 110958-19-5), PD-151832 (see Jaen etal. “In vitro and in vivo evaluation of the subtype-selective muscarinicagonist PD 151832.” Life Sci. 1995 56(11-12):845-52), vinconate (CAS RN70704-03-9), PYM-50028 PYM-50028 (Cogane) or PYM-50018 (Myogane) asdescribed by Harvey (“Natural Products in Drug Discovery andDevelopment. 27-28 Jun. 2005, London, UK.” IDrugs. 2005 8(9):719-21),SR-46559A (3-[N-(2 diethyl-amino-2-methylpropyl)-6-phenyl-5-propyl),dihydroergocristine (CAS RN 17479-19-5), dabelotine (CAS RN118976-38-8), zanapezil (CAS RN 142852-50-4).

Further non-limiting examples include NBI-113 (from NeurocrineBiosciences, Inc.), NDD-094 (from Novartis), P-58 or P58 (from Pfizer),or SR-57667 (from Sanofi-Synthelabo).

Moreover, an agent in combination with an angiotensin agent may be areported modulator of the nicotinic receptor. Non-limiting examples ofsuch a modulator include nicotine, acetylcholine, carbamylcholine,epibatidine, ABT-418 (structurally similar to nicotine, with anixoxazole moiety replacing the pyridyl group of nicotine), epiboxidine(a structural analogue with elements of both epibatidine and ABT-418),ABT-594 (azetidine analogue of epibatidine), lobeline, SSR-591813,represented by the following formula:

or SIB-1508 (altinicline).

In additional embodiments, an agent used in combination with anangiotensin agent is a reported aromatase inhibitor. Reported aromataseinhibitors include, but are not limited to, nonsteroidal or steroidalagents. Non-limiting examples of the former, which inhibit aromatase viathe heme prosthetic group, include anastrozole (Arimidex®), letrozole(Femara®), or vorozole (Rivisor®). Non-limiting examples of steroidalaromatase inhibitors AIs, which inactivate aromatase, include, but arenot limited to, exemestane (Aromasin®), androstenedione, or formestane(Lentaron®).

Additional non-limiting examples of a reported aromatase for use in acombination or method as disclosed herein include aminoglutethimide,4-androstene-3,6,17-trione (or “6-OXO”), or zoledronic acid or Zometa®(CAS RN 118072-93-8).

Further embodiments include a combination of an angiotensin agent and areported selective estrogen receptor modulator (SERM) may be used asdescribed herein. Non-limiting examples include tamoxifen, raloxifene,toremifene, clomifene, bazedoxifene, arzoxifene, or lasofoxifene.Additional non-limiting examples include a steroid antagonist or partialagonist, such as centchroman, clomiphene, or droloxifene.

In other embodiments, a combination of an angiotensin agent and areported cannabinoid receptor modulator may be used as described herein.Non-limiting examples include synthetic cannabinoids, endogenouscannabinoids, or natural cannabinoids. In some embodiments, the reportedcannabinoid receptor modulator is rimonabant (SR141716 or Acomplia),nabilone, levonantradol, marinol, or sativex (an extract containing bothTHC and CBD). Non-limiting examples of endogenous cannabinoids includearachidonyl ethanolamine (anandamide); analogs of anandamide, such asdocosatetraenylethanolamide or homo-γ-linoenylethanolamide; N-acylethanolamine signalling lipids, such as the noncannabimimeticpalmitoylethanolamine or oleoylethanolamine; or 2-arachidonyl glycerol.Non-limiting examples of natural cannabinoids includetetrahydrocannabinol (THC), cannabidiol (CBD), cannabinol (CBN),cannabigerol (CBG), cannabichromene (CBC), cannabicyclol (CBL),cannabivarol (CBV), tetrahydrocannabivarin (THCV), cannabidivarin(CBDV), cannabichromevarin (CBCV), cannabigerovarin (CBGV), orcannabigerol monoethyl ether (CBGM).

In yet further embodiments, an agent used in combination with anangiotensin agent is a reported FAAH (fatty acid amide hydrolase)inhibitor. Non-limiting examples of reported inhibitor agents includeURB597 (3′-carbamoyl-biphenyl-3-yl-cyclohexylcarbamate); CAY10401(1-oxazolo[4,5-b]pyridin-2-yl-9-octadecyn-1-one); OL-135(1-oxo-1[5-(2-pyridyl)-2-yl]-7-phenylheptane); anandamide (CAS RN94421-68-8); AA-5-HT (see Bisogno et al. “Arachidonoylserotonin andother novel inhibitors of fatty acid amide hydrolase.” Biochem BiophysRes Commun. 1998 248(3):515-22); 1-Octanesulfonyl fluoride; or O-2142 oranother arvanil derivative FAAH inhibitor as described by Di Marzo etal. (“A structure/activity relationship study on arvanil, anendocannabinoid and vanilloid hybrid.” J Pharmacol Exp Ther. 2002300(3):984-91).

Further non-limiting examples include SSR 411298 (from Sanofi-Aventis),JNJ28614118 (from Johnson & Johnson), or SSR 101010 (fromSanofi-Aventis).

In additional embodiments, an agent in combination with an angiotensinagent may be a reported modulator of nitric oxide function. Onenon-limiting example is sildenafil (Viagra®).

In additional embodiments, an agent in combination with an angiotensinagent may be a reported modulator of prolactin or a prolactin modulator.

In additional embodiments, an agent in combination with an angiotensinagent is a reported anti-viral agent, with ribavirin and amantadine asnon-limiting examples.

In additional embodiments, an agent in combination with an angiotensinagent may be a component of a natural product or a derivative of such acomponent. In some embodiments, the component or derivative thereof isin an isolated form, such as that which is separated from one or moremolecules or macromolecules normally found with the component orderivative before use in a combination or method as disclosed herein. Inother embodiments, the component or derivative is completely orpartially purified from one or more molecules or macromolecules normallyfound with the component or derivative. Exemplary cases of molecules ormacromolecules found with a component or derivative as described hereininclude a plant or plant part, an animal or animal part, and a food orbeverage product.

Non-limiting examples such a component include folic acid; a flavinoid,such as a citrus flavonoid; a flavonol, such as quercetin, kaempferol,myricetin, or isorhamnetin; a flavone, such as luteolin or apigenin; aflavanone, such as hesperetin, naringenin, or eriodictyol; a flavan-3-ol(including a monomeric, dimeric, or polymeric flavanol), such as(+)-catechin, (+)-gallocatechin, (−)-epicatechin, (−)-epigallocatechin,(−)-epicatechin 3-gallate, (−)-epigallocatechin 3-gallate, theaflavin,theaflavin 3-gallate, theaflavin 3′-gallate, theaflavin 3,3′ digallate,a thearubigin, or proanthocyanidin; an anthocyanidin, such as cyanidin,delphinidin, malvidin, pelargonidin, peonidin, or petunidin; anisoflavone, such as daidzein, genistein, or glycitein; flavopiridol; aprenylated chalcone, such as xanthohumol; a prenylated flavanone, suchas isoxanthohumol; a non-prenylated chalcone, such as chalconaringenin;a non-prenylated flavanone, such as naringenin; resveratrol; or ananti-oxidant neutraceutical (such as any present in chocolate, like darkchocolate or unprocessed or unrefined chocolate).

Additional non-limiting examples include a component of Gingko biloba,such as a flavo glycoside or a terpene. In some embodiments, thecomponent is a flavanoid, such as a flavonol or flavone glycoside, or aquercetin or kaempferol glycoside, or rutin; or a terpenoid, such asginkgolides A, B, C, or M, or bilobalide.

Further non-limiting examples include a component that is a flavanol, ora related oligomer, or a polyphenol as described in US2005/245601AA,US2002/018807AA, US2003/180406AA, US2002/086833AA, US2004/0236123,WO9809533, or WO9945788; a procyanidin or derivative thereof orpolyphenol as described in US2005/171029AA; a procyanidin, optionally incombination with L-arginine as described in US2003/104075AA; a low fatcocoa extract as described in US2005/031762AA; lipophilic bioactivecompound containing composition as described in US2002/107292AA; a cocoaextract, such as those containing one or more polyphenols orprocyanidins as described in US2002/004523AA; an extract of oxidized tealeaves as described in U.S. Pat. No. 5,139,802 or 5,130,154; a foodsupplement as described in WO 2002/024002.

Of course a composition comprising any of the above components, alone orin combination with an angiotensin agent as described herein is includedwithin the disclosure.

In additional embodiments, an agent in combination with an angiotensinagent may be a reported calcitonin receptor agonist such as calcitoninor the ‘orphan peptide’ PHM-27 (see Ma et al. “Discovery of novelpeptide/receptor interactions: identification of PHM-27 as a potentagonist of the human calcitonin receptor.” Biochem Pharmacol. 200467(7):1279-84). A further non-limiting example is the agonist fromKemia, Inc.

In an alternative embodiment, the agent may be a reported modulator ofparathyroid hormone activity, such as parathyroid hormone, or amodulator of the parathyroid hormone receptor.

In additional embodiments, an agent in combination with an angiotensinagent may a reported antioxidant, such as N-acetylcysteine oracetylcysteine; disufenton sodium (or CAS RN 168021-79-2 or Cerovive);activin (CAS RN 104625-48-1); selenium; L-methionine; an alpha, gamma,beta, or delta, or mixed, tocopherol; alpha lipoic acid; Coenzyme Q;benzimidazole; benzoic acid; dipyridamole; glucosamine; IRFI-016(2(2,3-dihydro-5-acetoxy-4,6,7-trimethylbenzofuranyl) acetic acid);L-carnosine; L-Histidine; glycine; flavocoxid (or LIMBREL®; baicalin,optionally with catechin (3,3′,4′,5,7-pentahydroxyflavan (2R,3S form)),and/or its stereo-isomer; masoprocol (CAS RN 27686-84-6); mesna (CAS RN19767-45-4); probucol (CAS RN 23288-49-5); silibinin (CAS RN22888-70-6); sorbinil (CAS RN 68367-52-2); spermine; tangeretin (CAS RN481-53-8); butylated hydroxyanisole (BHA); butylated hydroxytoluene(BHT); propyl gallate (PG); tertiary-butyl-hydroquinone (TBHQ);nordihydroguaiaretic acid (CAS RN 500-38-9); astaxanthin (CAS RN472-61-7); or an antioxidant flavonoid.

Additional non-limiting examples include a vitamin, such as vitamin A(Retinol) or C (Ascorbic acid) or E (including tocotrienol and/ortocopherol); a vitamin cofactors or mineral, such as coenzyme Q10(CoQ10), manganese, or melatonin; a carotenoid terpenoid, such aslycopene, lutein, alpha-carotene, beta-carotene, zeaxanthin,astaxanthin, or canthaxantin; a non-carotenoid terpenoid, such aseugenol; a flavonoid polyphenolic (or bioflavonoid); a flavonol, such asresveratrol, pterostilbene (methoxylated analogue of resveratrol),kaempferol, myricetin, isorhamnetin, a proanthocyanidin, or a tannin; aflavone, such as quercetin, rutin, luteolin, apigenin, or tangeritin; aflavanone, such as hesperetin or its metabolite hesperidin, naringeninor its precursor naringin, or eriodictyol; a flavan-3-ols(anthocyanidins), such as catechin, gallocatechin, epicatechin or agallate form thereof, epigallocatechin or a gallate form thereof,theaflavin or a gallate form thereof, or a thearubigin; an isoflavonephytoestrogens, such as genistein, daidzein, or glycitein; ananthocyanins, such as cyanidin, delphinidin, malvidin, pelargonidin,peonidin, or petunidin; a phenolic acid or ester thereof, such asellagic acid, gallic acid, salicylic acid, rosmarinic acid, cinnamicacid or a derivative thereof like ferulic acid, chlorogenic acid,chicoric acid, a gallotannin, or an ellagitannin; a nonflavonoidphenolic, such as curcumin; an anthoxanthin, betacyanin, citric acid,uric acid, R-α-lipoic acid, or silymarin.

Further non-limiting examples include 1-(carboxymethylthio)tetradecane;2,2,5,7,8-pentamethyl-1-hydroxychroman;2,2,6,6-tetramethyl-4-piperidinol-N-oxyl; 2,5-di-tert-butylhydroquinone;2-tert-butylhydroquinone; 3,4-dihydroxyphenylethanol; 3-hydroxypyridine;3-hydroxytamoxifen; 4-coumaric acid; 4-hydroxyanisole;4-hydroxyphenylethanol; 4-methylcatechol; 5,6,7,8-tetrahydrobiopterin;6,6′-methylenebis(2,2-dimethyl-4-methanesulfonicacid-1,2-dihydroquinoline);6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid;6-methyl-2-ethyl-3-hydroxypyridine; 6-O-palmitoylascorbic acid;acetovanillone; acteoside; actovegin; allicin; allyl sulfide;alpha-pentyl-3-(2-quinolinylmethoxy)benzenemethanol; alpha-tocopherolacetate; apolipoprotein A-IV; bemethyl; boldine; bucillamine; calciumcitrate; canthaxanthin; crocetin; diallyl trisulfide; dicarbine;dihydrolipoic acid; dimephosphon; ebselen; efamol; enkephalin-Leu,Ala(2)-Arg(6)-; ergothioneine; esculetin; essential 303 forte; ethonium;etofyllinclofibrate; fenozan; glaucine; H290-51; histidyl-prolinediketopiperazine; hydroquinone; hypotaurine; idebenone;indole-3-carbinol; isoascorbic acid; kojic acid, lacidipine, lodoxamidetromethamine; mexidol; morin; N,N′-diphenyl-4-phenylenediamine;N-isopropyl-N-phenyl-4-phenylenediamine; N-monoacetylcystine; nicaraven,nicotinoyl-GABA; nitecapone; nitroxyl; nobiletin; oxymethacil;p-tert-butyl catechol; phenidone; pramipexol; proanthocyanidin;procyanidin; prolinedithiocarbamate; propyl gallate; purpurogallin;pyrrolidine dithiocarbamic acid; rebamipide; retinol palmitate; salvin;selenious acid; sesamin; sesamol; sodium selenate; sodium thiosulfate;theaflavin; thiazolidine-4-carboxylic acid; tirilazad;tocopherylquinone; tocotrienol, alpha; a tocotrienol;tricyclodecane-9-yl-xanthogenate; turmeric extract; U 74389F; U 74500A;U 78517F; ubiquinone 9; vanillin; vinpocetine; xylometazoline; zetacarotene; zilascorb; zinc thionein; or zonisamide.

In additional embodiments, an agent in combination with an angiotensinagent may be a reported modulator of a norepinephrine receptor.Non-limiting examples include atomoxetine (Strattera®); a norepinephrinereuptake inhibitor, such as talsupram, tomoxetine, nortriptyline,nisoxetine, reboxetine (described, e.g., in U.S. Pat. No. 4,229,449), ortomoxetine (described, e.g., in U.S. Pat. No. 4,314,081); or a directagonist, such as a beta adrenergic agonist.

Additional non-limiting examples include an alpha adrenergic agonistsuch as etilefrine or a reported agonist of the α2-adrenergic receptor(or α2 adrenoceptor) like clonidine (CAS RN 4205-90-7), yohimbine,mirtazepine, atipamezole, carvedilol; dexmedetomidine or dexmedetomidinehydrochloride; ephedrine, epinephrine; etilefrine; lidamidine;tetramethylpyrazine; tizanidine or tizanidine hydrochloride;apraclonidine; bitolterol mesylate; brimonidine or brimonidine tartrate;dipivefrin (which is converted to epinephrine in vivo); guanabenz;guanfacine; methyldopa; alphamethylnoradrenaline; mivazerol; naturalephedrine or D(−)ephedrine; any one or any mixture of two, three, orfour of the optically active forms of ephedrine; CHF1035 or nolomirolehydrochloride (CAS RN 138531-51-8); or lofexidine (CAS RN 31036-80-3).

Alternative non-limiting examples include an adrenergic antagonist suchas a reported antagonist of the α2-adrenergic receptor like yohimbine(CAS RN 146-48-5) or yohimbine hydrochloride, idazoxan, fluparoxan,mirtazepine, atipamezole, or RX781094 (see Elliott et al. “Peripheralpre and postjunctional alpha 2-adrenoceptors in man: studies withRX781094, a selective alpha 2 antagonist.” J Hypertens Suppl. 19831(2):109-11).

Other non-limiting embodiments include a reported modulator of anα1-adrenergic receptor such as cirazoline; modafinil; ergotamine;metaraminol; methoxamine; midodrine (a prodrug which is metabolized tothe major metabolite desglymidodrine formed by deglycination ofmidodrine); oxymetazoline; phenylephrine; phenylpropanolamine; orpseudoephedrine.

Further non-limiting embodiments include a reported modulator of a betaadrenergic receptor such as arbutamine, befunolol, cimaterol,higenamine, isoxsuprine, methoxyphenamine, oxyfedrine, ractopamine,tretoquinol, or TQ-1016 (from TheraQuest Biosciences, LLC), or areported β1-adrenergic receptor modulator such as prenalterol, Ro 363,or xamoterol or a reported β1-adrenergic receptor agonist likedobutamine.

Alternatively, the reported modulator may be of a β2-adrenergic receptorsuch as levosalbutamol (CAS RN 34391-04-3), metaproterenol, MN-221 orKUR-1246((−)-bis(2-{[(2S)-2-({(2R)-2-hydroxy-2-[4-hydroxy-3-(2-hydroxyethyl)phenyl]ethyl}amino)-1,2,3,4-tetrahydronaphthalen-7-yl]oxy}-N,N-dimethylacetamide)monosulfateorbis(2-[[(2S)-2-([(2R)-2-hydroxy-2-[4-hydroxy-3-(2-hydroxyethyl)-phenyl]ethyl]amino)-1,2,3,4-tetrahydronaphthalen-7-yl]oxy]-N,N-dimethylacetamide)sulfateor CAS RN 194785-31-4), nylidrin, orciprenaline, pirbuterol, procaterol,reproterol, ritodrine, salmeterol, salmeterol xinafoate, terbutaline,tulobuterol, zinterol or bromoacetylalprenololmenthane, or a reportedP2-adrenergic receptor agonist like albuterol, albuterol sulfate,salbutamol (CAS RN 35763-26-9), clenbuterol, broxaterol, dopexamine,formoterol, formoterol fumarate, isoetharine, levalbuterol tartratehydrofluoroalkane, or mabuterol.

Additional non-limiting embodiments include a reported modulator of aβ3-adrenergic receptor such as AJ-9677 or TAK677([3-[(2R)-[[(2R)-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1H-indol-7-yloxy]aceticacid), or a reported β3-adrenergic receptor agonist like SR58611A(described in Simiand et al., Eur J Pharmacol, 219:193-201 (1992), BRL26830A, BRL 35135, BRL 37344, CL 316243 or ICI D7114.

Further alternative embodiments include a reported nonselective alphaand beta adrenergic receptor agonist such as epinephrine or ephedrine; areported nonselective alpha and beta adrenergic receptor antagonist suchas carvedilol; a β1 and β2 adrenergic receptor agonist such asisopreoterenol; or a β1 and β2 adrenergic receptor antagonist such asCGP 12177, fenoterol, or hexoprenaline.

Non-limiting examples of reported adrenergic agonists include albuterol,albuterol sulfate, salbutamol (CAS RN 35763-26-9), clenbuterol,adrafinil, and SR58611A (described in Simiand et al., Eur J Pharmacol,219:193-201 (1992)), clonidine (CAS RN 4205-90-7), yohimbine (CAS RN146-48-5) or yohimbine hydrochloride, arbutamine; befunolol; BRL 26830A;BRL 35135; BRL 37344; bromoacetylalprenololmenthane; broxaterol;carvedilol; CGP 12177; cimaterol; cirazoline; CL 316243; clenbuterol;denopamine; dexmedetomidine or dexmedetomidine hydrochloride;dobutamine, dopexamine, ephedrine, epinephrine, etilefrine; fenoterol;formoterol; formoterol fumarate; hexoprenaline; higenamine; ICI D7114;isoetharine; isoproterenol; isoxsuprine; levalbuterol tartratehydrofluoroalkane; lidamidine; mabuterol; methoxyphenamine; modafinil;nylidrin; orciprenaline; oxyfedrine; pirbuterol; prenalterol;procaterol; ractopamine; reproterol; ritodrine; ro 363; salmeterol;salmeterol xinafoate; terbutaline; tetramethylpyrazine; tizanidine ortizanidine hydrochloride; tretoquinol; tulobuterol; xamoterol; orzinterol. Additional non-limiting examples include apraclonidine,bitolterol mesylate, brimonidine or brimonidine tartrate, dipivefrin(which is converted to epinephrine in vivo), epinephrine, ergotamine,guanabenz, guanfacine, metaproterenol, metaraminol, methoxamine,methyldopa, midodrine (a prodrug which is metabolized to the majormetabolite desglymidodrine formed by deglycination of midodrine),oxymetazoline, phenylephrine, phenylpropanolamine, pseudoephedrine,alphamethylnoradrenaline, mivazerol, natural ephedrine or D(−)ephedrine,any one or any mixture of two, three, or four of the optically activeforms of ephedrine, CHF1035 or nolomirole hydrochloride (CAS RN138531-51-8), AJ-9677 or TAK677([3-[(2R)-[[(2R)-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1H-indol-7-yloxy]aceticacid), MN-221 or KUR-1246((−)-bis(2-{[(2S)-2-({(2R)-2-hydroxy-2-[4-hydroxy-3-(2-hydroxyethyl)phenyl]ethyl}amino)-1,2,3,4-tetrahydronaphthalen-7-yl]oxy}-N,N-dimethylacetamide)monosulfateorbis(2-[[(2S)-2-([(2R)-2-hydroxy-2-[4-hydroxy-3-(2-hydroxyethyl)-phenyl]ethyl]amino)-1,2,3,4-tetrahydronaphthalen-7-yl]oxy]-N,N-dimethylacetamide)sulfate or CAS RN 194785-31-4), levosalbutamol (CAS RN 34391-04-3),lofexidine (CAS RN 31036-80-3) or TQ-1016 (from TheraQuest Biosciences,LLC).

In further embodiments, a reported adrenergic antagonist, such asidazoxan or fluparoxan, may be used as an agent in combination with anootropic agent as described herein.

In further embodiments, an agent in combination with an angiotensinagent may be a reported modulator of carbonic anhydrase. Non-limitingexamples of such an agent include acetazolamide, benzenesulfonamide,benzolamide, brinzolamide, dichlorphenamide, dorzolamide or dorzolamideHCl, ethoxzolamide, flurbiprofen, mafenide, methazolamide, sezolamide,zonisamide, bendroflumethiazide, benzthiazide, chlorothiazide,cyclothiazide, dansylamide, diazoxide, ethinamate, furosemide,hydrochlorothiazide, hydroflumethiazide, mercuribenzoic acid,methyclothiazide, trichloromethazide, amlodipine, cyanamide, or abenzenesulfonamide. Additional non-limiting examples of such an agentinclude(4S-Trans)-4-(Ethylamino)-5,6-dihydro-6-methyl-4H-thieno(2,3-B)thiopyran-2-sulfonamide-7,7-dioxide;(4S-trans)-4-(methylamino)-5,6-dihydro-6-methyl-4H-thieno(2,3-B)thiopyran-2-sulfonamide-7,7-dioxide;(R)—N-(3-indol-1-yl-2-methyl-propyl)-4-sulfamoyl-benzamide;(S)—N-(3-indol-1-yl-2-methyl-propyl)-4-sulfamoyl-benzamide;1,2,4-triazole;1-methyl-3-oxo-1,3-dihydro-benzo[C]isothiazole-5-sulfonic acid amide;2,6-difluorobenzenesulfonamide; 3,5-difluorobenzenesulfonamide;3-mercuri-4-aminobenzenesulfonamide;3-nitro-4-(2-oxo-pyrrolidin-1-Yl)-benzenesulfonamide;4-(aminosulfonyl)-N-[(2,3,4-trifluorophenyl)methyl]-benzamide;4-(aminosulfonyl)-N-[(2,4,6-trifluorophenyl)methyl]-benzamide;4-(aminosulfonyl)-N-[(2,4-difluorophenyl)methyl]-benzamide;4-(aminosulfonyl)-N-[(2,5-difluorophenyl)methyl]-benzamide;4-(aminosulfonyl)-N-[(3,4,5-trifluorophenyl)methyl]-benzamide;4-(aminosulfonyl)-N-[(4-fluorophenyl)methyl]-benzamide;4-(hydroxymercury)benzoic acid; 4-fluorobenzenesulfonamide;4-methylimidazole; 4-sulfonamide-[1-(4-aminobutane)]benzamide;4-sulfonamide-[4-(thiomethylaminobutane)]benzamide;5-acetamido-1,3,4-thiadiazole-2-sulfonamide;6-oxo-8,9,10,11-tetrahydro-7H-cyclohepta[c][1]benzopyran-3-O-sulfamate;(4-sulfamoyl-phenyl)-thiocarbamic acid O-(2-thiophen-3-yl-ethyl)ester;(R)-4-ethylamino-3,4-dihydro-2-(2-methoylethyl)-2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;3,4-dihydro-4-hydroxy-2-(2-thienymethyl)-2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;3,4-dihydro-4-hydroxy-2-(4-methoxyphenyl)-2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;N-[(4-methoxyphenyl)methyl]2,5-thiophenedesulfonamide;2-(3-methoxyphenyl)-2H-thieno-[3,2-E]-1,2-thiazine-6-sulfinamide-1,1-dioxide;(R)-3,4-dihydro-2-(3-methoxyphenyl)-4-methylamino-2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;(S)-3,4-dihydro-2-(3-methoxyphenyl)-4-methylamino-2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;3,4-dihydro-2-(3-methoxyphenyl)-2H-thieno-[3,2-E]-1,2-thiazine-6-sulfonamide-1,1-dioxide;[2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide,2-(3-hydroxyphenyl)-3-(4-morpholinyl)-,1,1-dioxide];[2H-thieno[3,2-E]-1,2-thiazine-6-sulfonamide,2-(3-methoxyphenyl)-3-(4-morpholinyl)-,1,1-dioxide]; aminodi(ethyloxy)ethylaminocarbonylbenzenesulfonamide;N-(2,3,4,5,6-pentafluoro-benzyl)-4-sulfamoyl-benzamide;N-(2,6-difluoro-benzyl)-4-sulfamoyl-benzamide;N-(2-fluoro-benzyl)-4-sulfamoyl-benzamide;N-(2-thienylmethyl)-2,5-thiophenedisulfonamide;N-[2-(1H-indol-5-yl)-butyl]-4-sulfamoyl-benzamide;N-benzyl-4-sulfamoyl-benzamide; or sulfamic acid2,3-O-(1-methylethylidene)-4,5-O-sulfonyl-beta-fructopyranose ester.

In yet additional embodiments, an agent in combination with anangiotensin agent may be a reported modulator of acatechol-O-methyltransferase (COMT), such as floproprione, or a COMTinhibitor, such as tolcapone (CAS RN 134308-13-7), nitecapone (CAS RN116313-94-1), or entacapone (CAS RN 116314-67-1 or 130929-57-6).

In yet further embodiments, an agent in combination with an angiotensinagent may be a reported modulator of hedgehog pathway or signalingactivity such as cyclopamine, jervine, ezetimibe, regadenoson (CAS RN313348-27-5, or CVT-3146), a compound described in U.S. Pat. No.6,683,192 or identified as described in U.S. Pat. No. 7,060,450, orCUR-61414 or another compound described in U.S. Pat. No. 6,552,016.

In other embodiments, an agent in combination with an angiotensin agentmay be a reported modulator of IMPDH, such as mycophenolic acid ormycophenolate mofetil (CAS RN 128794-94-5).

In yet additional embodiments, an agent in combination with anangiotensin agent may be a reported modulator of a sigma receptor,including sigma-1 and sigma-2. Non-limiting examples of such a modulatorinclude an agonist of sigma-1 and/or sigma-2 receptor, such as(+)-pentazocine, SKF 10,047 (N-allylnormetazocine), or1,3-di-O-tolylguanidine (DTG). Additional non-limiting examples includeSPD-473 (from Shire Pharmaceuticals); a molecule with sigma modulatoryactivity as known in the field (see e.g., Bowen et al., PharmaceuticaActa Helvetiae 74: 211-218 (2000)); a guanidine derivative such as thosedescribed in U.S. Pat. Nos. 5,489,709; 6,147,063; 5,298,657; 6,087,346;5,574,070; 5,502,255; 4,709,094; 5,478,863; 5,385,946; 5,312,840; or5,093,525; WO9014067; an antipsychotic with activity at one or moresigma receptors, such as haloperidol, rimcazole, perphenazine,fluphenazine, (−)-butaclamol, acetophenazine, trifluoperazine,molindone, pimozide, thioridazine, chlorpromazine and triflupromazine,BMY 14802, BMY 13980, remoxipride, tiospirone, cinuperone (HR 375), orWY47384.

Additional non-limiting examples include igmesine; BD 1008 and relatedcompounds disclosed in U.S. Publication No. 20030171347; cis-isomers ofU50488 and related compounds described in de Costa et al, J. Med. Chem.,32(8): 1996-2002 (1989); U101958; SKF10,047; apomorphine; OPC-14523 andrelated compounds described in Oshiro et al., J Med Chem.; 43(2): 177-89(2000); arylcyclohexamines such as PCP; (+)-morphinans such asdextrallorphan; phenylpiperidines such as (+)-3-PPP and OHBQs;neurosteroids such as progesterone and desoxycorticosterone;butryophenones; BD614; or PRX-00023. Yet additional non-limitingexamples include a compound described in U.S. Pat. Nos. 6,908,914;6,872,716; 5,169,855; 5,561,135; 5,395,841; 4,929,734; 5,061,728;5,731,307; 5,086,054; 5,158,947; 5,116,995; 5,149,817; 5,109,002;5,162,341; 4,956,368; 4,831,031; or 4,957,916; U.S. Publication Nos.20050132429; 20050107432; 20050038011, 20030105079; 20030171355;20030212094; or 20040019060; European Patent Nos. EP 503 411; EP 362001-A1; or EP 461 986; International Publication Nos. WO 92/14464; WO93/09094; WO 92/22554; WO 95/15948; WO 92/18127; 91/06297; WO01/02380;WO91/18868; or WO 93/00313; or in Russell et al., J Med Chem.; 35(11):2025-33 (1992) or Chambers et al., J. Med Chem.; 35(11): 2033-9 (1992).

Further non-limiting examples include a sigma-1 agonist, such as IPAG(1-(4-iodophenyl)-3-(2-adamantyl)guanidine); pre-084; carbetapentane;4-IBP; L-687,384 and related compounds described in Middlemiss et al.,Br. J. Pharm., 102: 153 (1991); BD 737 and related compounds describedin Bowen et al., J Pharmacol Exp Ther., 262(1): 32-40 (1992)); OPC-14523or a related compound described in Oshiro et al., J Med Chem.; 43(2):177-89 (2000); a sigma-1 selective agonist, such as igmesine;(+)-benzomorphans, such as (+)-pentazocine and (+)-ethylketocyclazocine;SA-4503 or a related compound described in U.S. Pat. No. 5,736,546 or byMatsuno et al., Eur J Pharmacol., 306(1-3): 271-9 (1996); SK&F 10047; orifenprodil; a sigma-2 agonist, such as haloperidol,(+)-5,8-disubstituted morphan-7-ones, including CB 64D, CB 184, or arelated compound described in Bowen et al., Eur. J. Pharmacol.278:257-260 (1995) or Bertha et al., J. Med. Chem. 38:4776-4785 (1995);or a sigma-2 selective agonist, such as1-(4-fluorophenyl)-3-[4-[3-(4-fluorophenyl)-8-azabicyclo[3.2.1]oct-2-en-8-yl]-1-butyl]-1H-indole,Lu 28-179, Lu 29-253 or a related compound disclosed in U.S. Pat. No.5,665,725 or 6,844,352, U.S. Publication No. 20050171135, InternationalPatent Publication Nos. WO 92/22554 or WO 99/24436, Moltzen et al., J.Med Chem., 26; 38(11): 2009-17 (1995) or Perregaard et al., J Med Chem.,26; 38(11): 1998-2008 (1995).

Alternative non-limiting examples include a sigma-1 antagonist such asBD-1047(N(−)[2-(3,4-dichlorophenyl)ethyl]-N-methyl-2-(dimethylamin-o)ethylamine),BD-1063 (1(−)[2-(3,4-dichlorophenyl)ethyl]-4-methylpiperazine,rimcazole, haloperidol, BD-1047, BD-1063, BMY 14802, DuP 734, NE-100,AC915, or R-(+)-3-PPP. Particular non-limiting examples includefluoxetine, fluvoxamine, citalopram, sertaline, clorgyline, imipramine,igmesine, opipramol, siramesine, SL 82.0715, imcazole, DuP 734, BMY14802, SA 4503, OPC 14523, panamasine, or PRX-00023.

Other non-limiting examples of an agent in combination with anangiotensin agent include acamprosate (CAS RN 77337-76-9); a growthfactor, like LIF, EGF, FGF, bFGF or VEGF as non-limiting examples;octreotide (CAS RN 83150-76-9); an NMDA modulator like ketamine, DTG,(+)-pentazocine, DHEA, Lu 28-179(1′-[4-[1-(4-fluorophenyl)-1H-indol-3-yl]-1-butyl]-spiro[isobenzofuran-1(3H),4′-piperidine]), BD 1008 (CAS RN 138356-08-8), ACEA1021 (Licostinel orCAS RN 153504-81-5), GV150526A (Gavestinel or CAS RN 153436-22-7),sertraline, clorgyline, or memantine as non-limiting examples; ormetformin.

Additionally, the agent used with an angiotensin agent may be a reported5HT1a receptor agonist (or partial agonist) such as buspirone (buspar).In some embodiments, a reported 5HT1a receptor agonist is an azapirone,such as, but not limited to, tandospirone, gepirone and ipsapirone.Non-limiting examples of additional reported 5HT1a receptor agonistsinclude flesinoxan (CAS RN 98206-10-1), MDL 72832 hydrochloride,U-92016A, (+)-UH 301, F 13714, F 13640, 6-hydroxy-buspirone (see US2005/0137206), S-6-hydroxy-buspirone (see US 2003/0022899),R-6-hydroxy-buspirone (see US 2003/0009851), adatanserin,buspirone-saccharide (see WO 00/12067) or8-hydroxy-2-dipropylaminotetralin (8-OHDPAT).

Additional non-limiting examples of reported 5HT1a receptor agonistsinclude OPC-14523(1-[3-[4-(3-chlorophenyl)-1-piperazinyl]propyl]-5-methoxy-3,4-dihydro-2[1H]-quinolinonemonomethanesulfonate); BMS-181100 or BMY 14802 (CAS RN 105565-56-8);flibanserin (CAS RN 167933-07-5); repinotan (CAS RN 144980-29-0);lesopitron (CAS RN 132449-46-8); piclozotan (CAS RN 182415-09-4);Aripiprazole, Org-13011(1-(4-trifluoromethyl-2-pyridinyl)-4-[4-[2-oxo-1-pyrrolidinyl]butyl]piperazine(E)-2-butenedioate); SDZ-MAR-327 (see Christian et al. “Positronemission tomographic analysis of central dopamine D1 receptor binding innormal subjects treated with the atypical neuroleptic, SDZ MAR 327.” IntJ Mol. Med. 1998 1(1):243-7); MKC-242((S)-5-[3-[(1,4-benzodioxan-2-ylmethyl)amino]propoxy]-1,3-benzodioxoleHCl); vilazodone; sarizotan (CAS RN 177975-08-5); roxindole (CAS RN112192-04-8) or roxindole methanesulfonate (CAS RN 119742-13-1);alnespirone (CAS RN 138298-79-0); bromerguride (CAS RN 83455-48-5);xaliproden (CAS RN 135354-02-8); mazapertine succinate (CAS RN134208-18-7) or mazapertine (CAS RN 134208-17-6); PRX-00023; F-13640((3-chloro-4-fluoro-phenyl)-[4-fluoro-4-[[(5-methyl-pyridin-2-ylmethyl)-amino]methyl]piperidin-1-yl]methanone,fumaric acid salt); eptapirone (CAS RN 179756-85-5); Ziprasidone (CAS RN146939-27-7); Sunepitron (see Becker et al. “G protein-coupledreceptors: In silico drug discovery in 3D” PNAS 2004101(31):11304-11309); umespirone (CAS RN 107736-98-1); SLV-308;bifeprunox; and zalospirone (CAS RN 114298-18-9).

Yet further non-limiting examples include AP-521 (partial agonist fromAsahiKasei) and Du-123015 (from Solvay).

Alternatively, the agent used with an angiotensin agent may be areported 5HT4 receptor agonist (or partial agonist). In someembodiments, a reported 5HT4 receptor agonist or partial agonist is asubstituted benzamide, such as cisapride; individual, or a combinationof, cisapride enantiomers ((+) cisapride and (−) cisapride); mosapride;and renzapride as non-limiting examples. In other embodiments, thechemical entity is a benzofuran derivative, such as prucalopride.Additional embodiments include indoles, such as tegaserod, orbenzimidazolones. Other non-limiting chemical entities reported as a5HT4 receptor agonist or partial agonist include zacopride (CAS RN90182-92-6), SC-53116 (CAS RN 141196-99-8) and its racemate SC-49518(CAS RN 146388-57-0), BIMU1 (CAS RN 127595-43-1), TS-951 (CAS RN174486-39-6), or ML10302 CAS RN 148868-55-7). Additional non-limitingchemical entities include metoclopramide, 5-methoxytryptamine, RS67506,2-[1-(4-piperonyl)piperazinyl]benzothiazole, RS66331, BIMU8, SB 205149(the n-butyl quaternary analog of renzapride), or an indolecarbazimidamide as described by Buchheit et al. (“The serotonin 5-HT4receptor. 2. Structure-activity studies of the indole carbazimidamideclass of agonists.” J Med Chem. (1995) 38(13):2331-8). Yet additionalnon-limiting examples include norcisapride (CAS RN 102671-04-5) which isthe metabolite of cisapride; mosapride citrate; the maleate form oftegaserod (CAS RN 189188-57-6); zacopride hydrochloride (CAS RN99617-34-2); mezacopride (CAS RN 89613-77-4); SK-951((+−)-4-amino-N-(2-(1-azabicyclo(3.3.0)octan-5-yl)ethyl)-5-chloro-2,3-dihydro-2-methylbenzo[b]furan-7-carboxamidehemifumarate); ATI-7505, a cisapride analog from ARYx Therapeutics;SDZ-216-454, a selective 5HT4 receptor agonist that stimulates cAMPformation in a concentration dependent manner (see Markstein et al.“Pharmacological characterisation of 5-HT receptors positively coupledto adenylyl cyclase in the rat hippocampus.” Naunyn Schmiedebergs ArchPharmacol. (1999) 359(6):454-9); SC-54750, or Aminomethylazaadamantane;Y-36912, or4-amino-N-[1-[3-(benzylsulfonyl)propyl]piperidin-4-ylmethyl]-5-chloro-2-methoxybenzamideas disclosed by Sonda et al. (“Synthesis and pharmacological propertiesof benzamide derivatives as selective serotonin 4 receptor agonists.”Bioorg Med Chem. (2004) 12(10):2737-47); TKS159, or4-amino-5-chloro-2-methoxy-N-[(2S,4S)-1-ethyl-2-hydroxymethyl-4-pyrrolidinyl]benzamide,as reported by Haga et al. (“Effect of TKS159, a novel5-hydroxytryptamine-4 agonist, on gastric contractile activity inconscious dogs.”; RS67333, or1-(4-amino-5-chloro-2-methoxyphenyl)-3-(1-n-butyl-4-piperidinyl)-1-propanone;KDR-5169, or4-amino-5-chloro-N-[1-(3-fluoro-4-methoxybenzyl)piperidin-4-yl]-2-(2-hydroxyethoxy)benzamidehydrochloride dihydrate as reported by Tazawa, et al. (2002) “KDR-5169,a new gastrointestinal prokinetic agent, enhances gastric contractileand emptying activities in dogs and rats.” Eur J Pharmacol434(3):169-76); SL65.0155, or5-(8-amino-7-chloro-2,3-dihydro-1,4-benzodioxin-5-yl)-3-[1-(2-phenylethyl)-4-piperidinyl]-1,3,4-oxadiazol-2(3H)-one monohydrochloride; andY-34959, or4-Amino-5-chloro-2-methoxy-N-[1-[5-(1-methylindol-3-ylcarbonylamino)pentyl]piperidin-4-ylmethyl]benzamide.

Other non-limiting reported 5HT4 receptor agonists and partial agonistsfor use in combination with an angiotensin agent include metoclopramide(CAS RN 364-62-5), 5-methoxytryptamine (CAS RN 608-07-1), RS67506 (CASRN 168986-61-6), 2-[1-(4-piperonyl)piperazinyl]benzothiazole (CAS RN155106-73-3), RS66331 (see Buccafusco et al. “Multiple Central NervousSystem Targets for Eliciting Beneficial Effects on Memory andCognition.” (2000) Pharmacology 295(2):438-446), B1MU8(endo-N-8-methyl-8-azabicyclo[3.2.1]oct-3-yl)-2,3-dehydro-2-oxo-3-(prop-2-yl)-1H-benzimid-azole-1-carboxamide),or SB 205149 (the n-butyl quaternary analog of renzapride). Compoundsrelated to metoclopramide, such as metoclopramide dihydrochloride (CASRN 2576-84-3) or metoclopramide dihydrochloride (CAS RN 5581-45-3) ormetoclopramide hydrochloride (CAS RN 7232-21-5 or 54143-57-6) may alsobe used in a combination or method as described herein.

Additionally, the agent used with an angiotensin agent may be a reported5HT3 receptor antagonist such as azasetron (CAS RN 123039-99-6);Ondansetron (CAS RN 99614-02-5) or Ondansetron hydrochloride (CAS RN99614-01-4); Cilansetron (CAS RN 120635-74-7); Aloxi or PalonosetronHydrochloride (CAS RN 135729-62-3); Palenosetron (CAS RN 135729-61-2 or135729-56-5); Cisplatin (CAS RN 15663-27-1); Lotronex or Alosetronhydrochloride (CAS RN 122852-69-1); Anzemet or Dolasetron mesylate (CASRN 115956-13-3); zacopride or R-Zacopride; E-3620([3(S)-endo]-4-amino-5-chloro-N-(8-methyl-8-azabicyclo[3.2.1-]oct-3-yl-2-[(1-methyl-2-butynyl)oxy]benzamide)or E-3620HCl(3(S)-endo-4-amino-5-chloro-N-(8-methyl-8-azabicyclo[3.2.1]oct-3-yl)-2-(1-methyl-2-butinyl)oxy)-benzamide-HCl);YM 060 or Ramosetron hydrochloride (CAS RN 132907-72-3); athieno[2,3-d]pyrimidine derivative antagonist described in U.S. Pat. No.6,846,823, such as DDP 225 or MCI-225 (CAS RN 135991-48-9); Marinol orDronabinol (CAS RN 1972-08-3); or Lac Hydrin or Ammonium lactate (CAS RN515-98-0); Kytril or Granisetron hydrochloride (CAS RN 107007-99-8);Bemesetron (CAS RN 40796-97-2); Tropisetron (CAS RN 89565-68-4);Zatosetron (CAS RN 123482-22-4); Mirisetron (CAS RN 135905-89-4) orMirisetron maleate (CAS RN 148611-75-0); or renzapride (CAS RN112727-80-7).

Additionally, the agent used with an angiotensin agent may be a reported5HT2A/2C receptor antagonist such as Ketanserin (CAS RN 74050-98-9) orketanserin tartrate; risperidone; olanzapine; adatanserin (CAS RN127266-56-2); Ritanserin (CAS RN 87051-43-2); etoperidone; nefazodone;deramciclane (CAS RN 120444-71-5); Geoden or Ziprasidone hydrochloride(CAS RN 138982-67-9); Zeldox or Ziprasidone or Ziprasidonehydrochloride; EMD 281014(7-[4-[2-(4-fluoro-phenyl)-ethyl]-piperazine-1-carbonyl]-1H-indole-3-carbonitrileHCl); MDL 100907 or M100907 (CAS RN 139290-65-6); Effexor XR(Venlafaxine formulation); Zomaril or Iloperidone; quetiapine (CAS RN111974-69-7) or Quetiapine fumarate (CAS RN 111974-72-2) or Seroquel; SB228357 or SB 243213 (see Bromidge et al. “Biarylcarbamoylindolines arenovel and selective 5-HT(2C) receptor inverse agonists: identificationof5-methyl-1-[[2-[(2-methyl-3-pyridyl)oxy]-5-pyridyl]carbamoyl]-6-trifluoromethylindoline(SB-243213) as a potential antidepressant/anxiolytic agent.” J Med Chem.2000 43(6): 1123-34; SB 220453 or Tonabersat (CAS RN 175013-84-0);Sertindole (CAS RN 106516-24-9); Eplivanserin (CAS RN 130579-75-8) orEplivanserin fumarate (CAS RN 130580-02-8); Lubazodone hydrochloride(CAS RN 161178-10-5); Cyproheptadine (CAS RN 129-03-3); Pizotyline orpizotifen (CAS RN 15574-96-6); Mesulergine (CAS RN 64795-35-3);Irindalone (CAS RN 96478-43-2); MDL 11939 (CAS RN 107703-78-6); orpruvanserin (CAS RN 443144-26-1).

Additional non-limiting examples of modulators include reported 5-HT2Cagonists or partial agonists, such as m-chlorophenylpiperazine; or5-HT2A receptor inverse agonists, such as ACP 103 (CAS RN: 868855-07-6),APD125 (from Arena Pharmaceuticals), AVE 8488 (from Sanofi-Aventis) orTGWOOAD/AA (from Fabre Kramer Pharmaceuticals).

Additionally, the agent used with an angiotensin agent may be a reported5HT6 receptor antagonist such as SB-357134(N-(2,5-Dibromo-3-fluorophenyl)-4-methoxy-3-piperazin-1-ylbenzenesulfonamide);SB-271046(5-chloro-N-(4-methoxy-3-(piperazin-1-yl)phenyl)-3-methylbenzo[b]thiophene-2-sulfonamide);Ro 04-06790(N-(2,6-bis(methylamino)pyrimidin-4-yl)-4-aminobenzenesulfonamide); Ro63-0563 (4-amino-N-(2,6 bis-methylamino-pyridin-4-yl)-benzenesulfonamide); clozapine or its metabolite N-desmethylclozapine;olanzapine (CAS RN 132539-06-1); fluperlapine (CAS RN 67121-76-0);seroquel (quetiapine or quetiapine fumarate); clomipramine (CAS RN303-49-1); amitriptyline (CAS RN50-48-6); doxepin (CAS RN 1668-19-5);nortryptyline (CAS RN 72-69-5); 5-methoxytryptamine (CAS RN 608-07-1);bromocryptine (CAS RN 25614-03-3); octoclothepin (CAS RN 13448-22-1);chlorpromazine (CAS RN 50-53-3); loxapine (CAS RN 1977-10-2);fluphenazine (CAS RN 69-23-8); or GSK 742457 (presented by David Witty,“Early Optimisation of in vivo Activity: the discovery of 5-HT6 ReceptorAntagonist 742457” GlaxoSmithKline at SCIpharm 2006, InternationalPharmaceutical Industry Conference in Edinburgh, 16 May 2006).

As an additional non-limiting example, the reported 5HT6 modulator maybe SB-258585(4-Iodo-N-[4-methoxy-3-(4-methyl-piperazin-1-yl)-phenyl]-benzenesulphonamide);PRX 07034 (from Predix Pharmaceuticals) or a partial agonist, such asE-6801(6-chloro-N-(3-(2-(dimethylamino)ethyl)-1H-indol-5-yl)imidazo[2,1-b]thiazole-5-sulfonamide)or E-6837(5-chloro-N-(3-(2-(dimethylamino)ethyl)-1H-indol-5-yl)naphthalene-2-sulfonamide).

In additional embodiments, the neurogenic agent is ethyleicosapentaenoate or ethyl-EPA (also known as5,8,11,14,17-eicosapentaenoic acid ethyl ester or miraxion, ChemicalAbstracts Registry number 86227-47-6), docosahexaenoic acid (DHA), or aretinoid acid drug.

Having now generally described the invention, the same will be morereadily understood through reference to the following examples which areprovided by way of illustration, and are not intended to be limiting ofthe present invention, unless specified.

EXAMPLES Example 1 Effect of Alacepril on Neuronal Differentiation ofHuman Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of alacepril (testcompound), and stained with TUJ-1 antibody, as described in PCTApplication No. US06/026677 (incorporated by reference). Mitogen-freetest media with a positive control for neuronal differentiation was usedalong with basal media without growth factors as a negative control.

Results are shown in FIG. 1, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that alacepril promoted neuronaldifferentiation.

Example 2 Effect of Enalapril on Neuronal Differentiation of HumanNeural Stem Cells

hNSCs were prepared and treated with varying concentrations of enalapril(test compound), and stained with TUJ-1 antibody, as described inExample 1. Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 2, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that enalapril promoted neuronaldifferentiation.

Example 3 Effect of Lisinopril on Neuronal Differentiation of HumanNeural Stem Cells

hNSCs were prepared and treated with varying concentrations oflisinopril (test compound), and stained with TUJ-1 antibody, asdescribed in Example 1. Mitogen-free test media with a positive controlfor neuronal differentiation was used along with basal media withoutgrowth factors as a negative control.

Results are shown in FIG. 3, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that lisinopril promoted neuronaldifferentiation.

Example 4 Effect of Captopril on Neuronal Differentiation of HumanNeural Stem Cells

hNSCs were prepared and treated with varying concentrations of captopril(test compound), and stained with TUJ-1 antibody, as described inExample 1. Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 4, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that captopril promoted neuronaldifferentiation.

Example 5 Effect of Losartan on Neuronal Differentiation of Human NeuralStem Cells

hNSCs were prepared and treated with varying concentrations of losartan(test compound), and stained with TUJ-1 antibody, as described inExample 1. Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 5, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that losartan promoted neuronaldifferentiation.

Example 6 Effect of Benazepril on Neuronal Differentiation of HumanNeural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of benazepril (testcompound), and stained with TUJ-1 antibody, as described above.Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 6, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that benazepril promoted neuronaldifferentiation.

Example 7 Effect of Trandolapril on Neuronal Differentiation of HumanNeural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of trandolapril(test compound), and stained with TUJ-1 antibody, as described in above.Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 7, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that trandolapril promoted neuronaldifferentiation.

Example 8 Effect of Candesartan on Neuronal Differentiation of HumanNeural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of candesartan(test compound), and stained with TUJ-1 antibody, as described in above.Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 8, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that candesartan promoted neuronaldifferentiation.

Example 9 Effect of Telmisartan on Neuronal Differentiation of HumanNeural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of telmisartan(test compound), and stained with TUJ-1 antibody, as described in above.Mitogen-free test media with a positive control for neuronaldifferentiation was used along with basal media without growth factorsas a negative control.

Results are shown in FIG. 9, which shows dose response curves ofneuronal differentiation after background media values are subtracted.The dose response curve of the neuronal positive control is included asa reference. The data is presented as a percent of neuronal positivecontrol. The data indicate that telmisartan promoted neuronaldifferentiation.

Example 10 Effect of Combining Captopril and Ibudilast on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of ibudilast and/orcaptopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 10, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofibudilast and captopril is shown with the concentration response curvesof ibudilast or captopril alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofibudilast with captopril resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 11 Effect of Combining Captopril and Enoximone on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of enoximone and/orcaptopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 11, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofenoximone and captopril is shown with the concentration response curvesof enoximone or captopril alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofenoximone with captopril resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 12 Effect of Combining Ibudilast and Candesartan on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of ibudilast and/orcandesartan (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 12, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofibudilast and candesartan is shown with the concentration responsecurves of ibudilast or candesartan alone. The data is presented as apercent of neuronal positive control. The data indicate that thecombination of ibudilast with candesartan resulted in synergisticallyenhanced neuronal differentiation relative to that that produced byeither agent alone.

Example 13 Effect of Combining Captopril and Baclofen on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of baclofen and/orcaptopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 13, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofbaclofen and captopril is shown with the concentration response curvesof baclofen or captopril alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofbaclofen with captopril resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 14 Effect of Combining Captopril and Donepezil on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of donepezil and/orcaptopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 14, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofdonepezil and captopril is shown with the concentration response curvesof donepezil or captopril alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofdonepezil with captopril resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 15 Effect of Combining Captopril and Vardenafil on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of vardenafiland/or captopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 15, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofvardenafil and captopril is shown with the concentration response curvesof vardenafil or captopril alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofvardenafil with captopril resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 16 Effect of Combining Telmisartan and Rolipram on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of rolipram and/ortelmisartan (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 16, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination ofrolipram and telmisartan is shown with the concentration response curvesof rolipram or telmisartan alone. The data is presented as a percent ofneuronal positive control. The data indicate that the combination ofrolipram with telmisartan resulted in synergistically enhanced neuronaldifferentiation relative to that that produced by either agent alone.

Example 17 Determination of Synergy

The presence of synergy was determined by use of a combination index(CI). The CI based on the EC₅₀ was used to determine whether a pair ofcompounds had an additive, synergistic (greater than additive), orantagonistic effect when run in combination. The CI is a quantitativemeasure of the nature of drug interactions, comparing the EC₅₀'s of twocompounds, when each is assayed alone, to the EC₅₀ of each compound whenassayed in combination. The combination index (CI) is equal to thefollowing formula:

$\frac{C\; 1}{{IC}\; 1} + \frac{C\; 2}{{IC}\; 2} + \frac{\left( {C\; 1*C\; 2} \right)}{\left( {{IC}\; 1*{IC}\; 2} \right)}$

where C1 and C2 are the concentrations of a first and a second compound,respectively, resulting in 50% activity in neuronal differentiation whenassayed in combination; and IC1 and IC2 are the concentrations of eachcompound resulting in 50% activity when assayed independently. A CI ofless than 1 indicates the presence of synergy; a CI equal to 1 indicatesan additive effect; and a CI greater than 1 indicates antagonism betweenthe two compounds.

Non-limiting examples of combinations of an angiotensin agent and anadditional agent as described herein were observed to result insynergistic activity. The exemplary results are shown in the followingtable:

Combination CI Captopril + Ibudilast 0.05 Captopril + Enoximone 0.5Candesartan + Ibudilast 0.07 Captopril + Baclofen 0.88 Captopril +Donepezil 0.13 Captopril + Vardenafil 0.69 Telmisartan + Rolipram 0.62Captopril + Theophylline 0.07

As the CI is less than 1 for each of these combinations, the twocompounds have a synergistic effect in neuronal differentiation.

The above is based on the selection of EC₅₀ as the point of comparisonfor the two compounds. The comparison is not limited by the point used,but rather the same comparison may be made at another point, such asEC₂₀, EC₃₀, EC₄₀, EC₆₀, EC₇₀, EC₈₀, or any other EC value above, below,or between any of those points.

Example 18 Effect of Combining Captopril and Theophylline on NeuronalDifferentiation of Human Neural Stem Cells

Human neural stem cells (hNSCs) were isolated and grown in monolayerculture, plated, treated with varying concentrations of theophyllineand/or captopril (test compounds), and stained with TUJ-1 antibody, asdescribed above. Mitogen-free test media with a positive control forneuronal differentiation was used along with basal media without growthfactors as a negative control.

Results are shown in FIG. 17, which shows concentration response curvesof neuronal differentiation after background media values aresubtracted. The concentration response curve of the combination oftheophylline and captopril is shown with the concentration responsecurves of theophylline or captopril alone. The data is presented as apercent of neuronal positive control. The data indicate that thecombination of theophylline with captopril resulted in synergisticallyenhanced neuronal differentiation relative to that that produced byeither agent alone.

All references cited herein, including patents, patent applications, andpublications, are hereby incorporated by reference in their entireties,whether previously specifically incorporated or not.

Having now fully described this invention, it will be appreciated bythose skilled in the art that the same can be performed within a widerange of equivalent parameters, concentrations, and conditions withoutdeparting from the spirit and scope of the invention and without undueexperimentation.

While this invention has been described in connection with specificembodiments thereof, it will be understood that it is capable of furthermodifications. This application is intended to cover any variations,uses, or adaptations of the invention following, in general, theprinciples of the invention and including such departures from thepresent disclosure as come within known or customary practice within theart to which the invention pertains and as may be applied to theessential features hereinbefore set forth.

1. A composition comprising, captopril in combination with or donepezil.2. The composition of claim 1, wherein the combination is in a singleformulation.